JP2020011125A - Autoassembling peptides for treatment of pulmonary leakage - Google Patents

Abstract

To provide materials and methods for treatment of pulmonary leakage.SOLUTION: A peptide comprising about 7 to about 32 amino acids in a solution may be introduced to a target site. A hydrogel barrier may be provided at the target site in order to treat the pulmonary leakage. The method also includes administering through a delivery device a solution comprising a self-assembling peptide comprising about 7-32 amino acids in an effective amount and in an effective concentration to the target area to form a hydrogel barrier under physiological conditions of the target area in order to treat the pulmonary leakage.SELECTED DRAWING: None

Description

SEQUENCE LISTING This application contains a sequence listing filed electronically in ASCII format and incorporated herein by reference in its entirety. The ASCII copy created on March 10, 2015 is T2071-7006 WO_SL. txt, and the size is 1,751 bytes.

  The present disclosure relates generally to materials and methods that can be used in medical, research, and industrial applications. More specifically, the present disclosure relates to materials and methods that can be used to treat pulmonary leaks, including, for example, pleural defects and pleural leaks from bronchial anastomosis leaks.

In embodiments, a method is provided for treating lung leakage in a subject. The method includes introducing a delivery device to a leak targeted area of the subject's lungs. The method also includes positioning the distal end of the delivery device within a target area where treatment of a lung leak is desired. The method also comprises administering to the target area a solution comprising an effective amount and concentration of a self-assembling peptide comprising about 7 to 32 amino acids through a delivery device to treat lung leakage. Forming a hydrogel barrier under physiological conditions. The method also includes removing the delivery device from the target area.
The present invention provides, for example, the following items.
(Item 1)
A method of treating lung leakage in a subject, comprising:
Introducing a delivery device to a target area of the lung leak of the subject;
Placing the distal end of the delivery device within the target area where treatment of lung leakage is desired;
Administering a solution comprising an effective amount and an effective concentration of a self-assembling peptide comprising from about 7 amino acids to 32 amino acids through the delivery device to the target area to treat the lung leakage; Forming a hydrogel barrier underneath;
Removing the delivery device from the target area.
(Item 2)
The method of claim 1, further comprising, before the step of introducing the delivery device, visualizing a region including the target area.
(Item 3)
The method of claim 1, further comprising, after removing the delivery device from the target area, visualizing an area including the target area.
(Item 4)
The method of claim 1, further comprising, after the step of removing the delivery device, monitoring the target area.
(Item 5)
The method of claim 1, wherein administering the solution comprises applying the solution locally to the target area.
(Item 6)
The method of claim 1, wherein administering the solution comprises injecting the solution into the target area by overflow to cover the target area locally.
(Item 7)
2. The method of item 1, wherein administering the solution comprises administering the solution in a single dose.
(Item 8)
2. The method of item 1, wherein administering the solution comprises administering the solution in at least two doses.
(Item 9)
The hydrogel barrier, resulting in rupture pressure resistant of at least 35 cm H ₂ O, The method of claim 1.
(Item 10)
The method of claim 1, wherein the hydrogel barrier is formed in less than about 3 minutes.
(Item 11)
11. The method according to item 10, wherein the hydrogel barrier is formed in less than about 2 minutes.
(Item 12)
12. The method according to item 11, wherein the hydrogel barrier is formed in less than about 1 minute.
(Item 13)
The method of claim 1, wherein the hydrogel barrier is formed between about 2 seconds and about 30 seconds.
(Item 14)
2. The method according to item 1, further comprising preparing the solution containing the self-assembling peptide.
(Item 15)
2. The method of item 1, further comprising evaluating the subject to determine a need to treat lung leakage and preparing the solution based on the evaluating.
(Item 16)
Item 16. The method according to item 14 or 15, further comprising adjusting a pH of the solution. (Item 17)
Item 16. The method according to item 14 or 15, further comprising increasing the pH of the solution.
(Item 18)
2. The method of claim 1, wherein at least one of the effective amount and the effective concentration is based in part on a size of the target area of leakage of the lung.
(Item 19)
19. The method according to item 18, wherein the effective amount is approximately 1 mL per cm ^{2 of the} target area. (Item 20)
19. The method of item 18, wherein the amount effective to enable treatment of lung leakage comprises a volume in the range of about 0.1 mL to about 5 mL.
(Item 21)
2. The method according to item 1, wherein the solution is substantially free of cells.
(Item 22)
2. The method according to item 1, wherein the solution is substantially free of a drug.
(Item 23)
The method of claim 1, further comprising administering the solution after a surgical procedure.
(Item 24)
24. The method of claim 23, wherein the surgical procedure resulted in leakage of the lung.
(Item 25)
2. The method according to item 1, wherein the lung leakage is a pleural defect.
(Item 26)
The method of claim 1, wherein the lung leak is a bronchial anastomosis leak.
(Item 27)
2. The self-assembling peptide according to item 1, wherein the self-assembling peptide is selected from the group consisting of (RADA) ₄ (SEQ ID NO: 1), (IEIK) ₃ I (SEQ ID NO: 2), and (KLDL) ₃ (SEQ ID NO: 3). the method of.
(Item 28)
Item 27, wherein the concentration effective to allow treatment of the lung leakage comprises a self-assembling peptide concentration ranging from about 0.1 weight / volume (w / v) percent to about 3 w / v percent. The described method.
(Item 29)
28. The method of item 27, wherein preparing the solution containing the self-assembling peptide comprises adding the self-assembling peptide to a saline solution.
(Item 30)
Preparing the solution containing the self-assembling peptide,
Providing an aqueous peptide solution by adding water to the peptide powder of the self-assembling peptide,
Adding a saline solution to the aqueous peptide solution,
28. The method according to item 27, comprising mixing the salt solution and the aqueous peptide solution.
(Item 31)
Wherein the salt solution comprises at least one cation selected from the group consisting of ammonium ion, iron ion, magnesium ion, potassium ion, pyridinium ion, quaternary ammonium ion, sodium ion, potassium ion, and calcium ion, 31. The method according to item 29 or 30.
(Item 32)
The salt solution is composed of chloride ion, sulfate ion, acetate ion, carbonate ion, chloride ion, citrate ion, cyanide ion, fluoride ion, sulfate ion, nitrate ion, nitrite ion, and phosphate ion. 31. The method according to item 29 or 30, comprising at least one anion selected from the group.
(Item 33)
32. The method according to item 31, wherein the salt solution includes at least one of calcium chloride, sodium chloride, and potassium chloride.
(Item 34)
29. The method according to item 28, wherein the solution comprising the self-assembling peptide comprises (RADA) ₄ (SEQ ID NO: 1) at a concentration of about 0.5 weight / volume (w / v) percent.
(Item 35)
35. The method according to item 34, wherein the solution containing the self-assembling peptide comprises a calcium chloride concentration of about 0.125M.
(Item 36)
36. The method according to item 35, wherein the solution containing the self-assembling peptide has a storage modulus of about 25 Pa.
(Item 37)
35. The method of claim 34, wherein said solution comprising said self-assembling peptide comprises a calcium chloride concentration of about 0.250M.
(Item 38)
38. The method according to item 37, wherein the solution containing the self-assembling peptide has a storage modulus of about 44 Pa.
(Item 39)
35. The method of claim 34, wherein said solution comprising said self-assembling peptide comprises a calcium chloride concentration of about 0.500M.
(Item 40)
40. The method according to item 39, wherein the solution containing the self-assembling peptide has a storage modulus of about 52 Pa.
(Item 41)
28. The method of claim 27, wherein the solution comprising the self-assembling peptide comprises (RADA) ₄ (SEQ ID NO: 1) at a concentration of about 2.5 weight / volume (w / v) percent.
(Item 42)
42. The method according to item 41, wherein the solution containing the self-assembling peptide comprises a calcium chloride concentration of about 0.125M.
(Item 43)
43. The method according to item 42, wherein the solution containing the self-assembling peptide has a storage modulus of about 600 Pa.
(Item 44)
29. The method according to item 28, wherein the solution containing the self-assembling peptide has a salt concentration of about 0.005M to about 1M.
(Item 45)
45. The method of item 44, wherein the solution comprising the self-assembling peptide has a salt concentration between about 0.125M and about 0.500M.
(Item 46)
46. The method according to item 45, wherein the solution containing the self-assembling peptide has a salt concentration of about 0.25M.
(Item 47)
29. The method according to item 28, further comprising a solution comprising sodium chloride, potassium chloride, calcium chloride, and sodium bicarbonate.
(Item 48)
29. The method according to item 28, further comprising a solution containing a contrast agent.
(Item 49)
49. The method according to item 48, wherein the contrast agent includes a sulfate ion and a sodium ion.
(Item 50)
29. The method according to item 28, wherein the solution has a pH of about 2.5 to about 4.0.
(Item 51)
Item 50, wherein the solution has a pH of about 3.5 and the self-assembling peptide is one of (RADA) ₄ (SEQ ID NO: 1) and (KLDL) ₃ (SEQ ID NO: 3). the method of.
(Item 52)
Wherein the solution has a pH of about 3.7, the self-assembling peptide is a _(IEIK) 3 I (SEQ ID NO: 2) The method of claim 50.
(Item 53)
16. The method of claim 14, wherein preparing a solution comprising the self-assembling peptide comprises one of adding the self-assembling peptide to a buffer and adding a buffer to the solution. Method.
(Item 54)
54. The method according to item 53, wherein the buffer comprises at least two salts.
(Item 55)
55. The method according to item 54, wherein the buffer has a pH of 7.2.
(Item 56)
55. The method according to item 54, wherein the buffer has a pH of 7.4.
(Item 57)
54. The method according to item 53, wherein the buffer is an alkaline buffer.
(Item 58)
54. The method of claim 53, wherein said solution is buffered with at least one of about 0.15 M sodium chloride, potassium chloride, magnesium chloride, and calcium chloride.
(Item 59)
59. The method of claim 58, wherein said buffer comprises between about 0.6 M to about 1.2 M salt, and wherein said self-assembling peptide is (RADA) ₄ (SEQ ID NO: 1).
(Item 60)
Said buffer comprises a salt of between about 0.02M to about 0.04 M, the a self-assembled peptide _(IEIK) 3 I (SEQ ID NO: 2) The method of claim 58.
(Item 61)
59. The method of claim 58, wherein said buffer comprises between about 0.1 M to about 0.4 M salt, and wherein said self-assembling peptide is (KLDL) ₃ (SEQ ID NO: 3).
(Item 62)
29. The method according to item 28, further comprising selecting a salt to provide a predetermined mechanical strength to the solution.
(Item 63)
63. The method of claim 62, further comprising selecting the concentration of the salt to provide the predetermined mechanical strength to the solution.
(Item 64)
29. The method of item 28, further comprising selecting a salt to provide a predetermined ionic strength to the solution.
(Item 65)
65. The method of claim 64, further comprising selecting a concentration of the salt such that the solution has the predetermined ionic strength.
(Item 66)
29. The method according to item 28, further comprising selecting a salt to bring a predetermined pH to the solution.
(Item 67)
67. The method of claim 66, further comprising selecting a concentration of the salt such that the solution has the predetermined pH.
(Item 68)
2. The method according to item 1, wherein the subject is a mammal.
(Item 69)
69. The method according to item 68, wherein the subject is a human.
(Item 70)
2. The method according to item 1, wherein the self-assembling peptide contains about 12 to about 16 amino acids in which hydrophobic amino acids and hydrophilic amino acids alternate.
(Item 71)
2. The method of claim 1, wherein said solution further comprises at least one biologically active agent.
(Item 72)
A kit for treating lung leakage in a subject, comprising:
A self-assembling peptide comprising an effective amount of about 7 amino acids to about 32 amino acids to form a hydrogel barrier under physiological conditions to enable treatment of lung leakage;
Instructions for administering the self-assembling peptide to a target area of leakage of the subject's lungs.
(Item 73)
73. The kit according to item 72, wherein the self-assembling peptide is provided as one of a solution containing the self-assembling peptide and a powder prepared as a solution containing the self-assembling peptide.
(Item 74)
74. The kit according to item 73, wherein the self-assembling peptide is provided as a solution containing the self-assembling peptide.
(Item 75)
74. The kit according to item 73, wherein the self-assembling peptide is provided as a powder prepared as a solution containing the self-assembling peptide.
(Item 76)
74. The method of claim 73, further comprising instructions for preparing a solution comprising a self-assembling peptide having an effective concentration to form a hydrogel barrier under physiological conditions to enable treatment of the lung leakage. Kit.
(Item 77)
73. The kit of claim 72, further comprising a delivery device for introducing the self-assembling peptide to a target area of the lung leak.
(Item 78)
73. The kit according to item 72, wherein the lung leakage is a pleural defect.
(Item 79)
73. The kit according to item 72, wherein the leak of the lung is a leak of bronchial anastomosis.
(Item 80)
Item 73, wherein the self-assembling peptide is selected from the group consisting of (RADA) ₄ (SEQ ID NO: 1), (IEIK) ₃ I (SEQ ID NO: 2), and (KLDL) ₃ (SEQ ID NO: 3). Kit.
(Item 81)
Item 80, wherein the effective concentration to enable treatment of the lung leak comprises a concentration of the self-assembling peptide ranging from about 0.1 weight / volume (w / v) percent to about 3 w / v percent. The kit according to 1.
(Item 82)
73. The kit according to item 72, further comprising a saline solution.
(Item 83)
83. The kit of item 82, further comprising instructions for combining the saline solution with one of a solution comprising the self-assembling peptide and a peptide powder.
(Item 84)
Wherein the salt solution comprises at least one cation selected from the group consisting of ammonium ion, iron ion, magnesium ion, potassium ion, pyridinium ion, quaternary ammonium ion, sodium ion, potassium ion, and calcium ion, 89. The kit according to item 82.
(Item 85)
The salt solution is composed of chloride ion, sulfate ion, acetate ion, carbonate ion, chloride ion, citrate ion, cyanide ion, fluoride ion, sulfate ion, nitrate ion, nitrite ion, and phosphate ion. 83. The kit according to item 82, comprising at least one anion selected from the group.
(Item 86)
85. The kit according to item 84, wherein the salt solution comprises at least one of calcium chloride, sodium chloride, and potassium chloride.
(Item 87)
85. The kit according to item 84, wherein the solution comprising the self-assembling peptide comprises a salt concentration of between about 0.005M and about 0.500M.
(Item 88)
90. The kit according to item 87, wherein the solution comprising the self-assembling peptide has a storage modulus between about 25 Pa and about 600 Pa.
(Item 89)
90. The kit according to item 87, wherein the solution containing the self-assembling peptide has a salt concentration of about 0.25M.
(Item 90)
73. The kit of item 72, further comprising a solution comprising sodium chloride, potassium chloride, calcium chloride, and sodium bicarbonate.
(Item 91)
73. The kit according to item 72, further comprising a solution containing a contrast agent.
(Item 92)
90. The kit according to item 91, wherein the contrast agent comprises a sulfate ion and a sodium ion.
(Item 93)
73. The kit according to item 72, wherein the solution containing the self-assembling peptide has a pH of about 2.5 to about 4.0.
(Item 94)
The solution comprising the self-assembling peptide has a pH of about 3.5, and the self-assembling peptide is one of (RADA) ₄ (SEQ ID NO: 1) and (KLDL) ₃ (SEQ ID NO: 3) 100. The kit according to item 93, wherein
(Item 95)
94. The kit of item 93, wherein the solution comprising the self-assembling peptide has a pH of about 3.7, and wherein the self-assembling peptide is (IEIK) ₃ I (SEQ ID NO: 2).
(Item 96)
73. The kit according to item 72, wherein one of the kit or the solution containing the self-assembling peptide comprises a buffer.
(Item 97)
97. The kit according to item 96, wherein the buffer comprises at least two salts.
(Item 98)
97. The kit according to item 96, wherein the buffer has a pH of 7.2.
(Item 99)
97. The kit according to item 96, wherein the buffer has a pH of 7.4.
(Item 100)
97. The kit according to item 96, wherein the buffer is an alkaline buffer.
(Item 101)
97. The kit according to item 96, wherein the solution is buffered with at least one of about 0.15 M sodium chloride, potassium chloride, magnesium chloride, and calcium chloride.
(Item 102)
101. The kit of item 101, wherein said buffer comprises between about 0.6M to about 1.2M salt, and wherein said self-assembling peptide is (RADA) ₄ (SEQ ID NO: 1).
(Item 103)
The buffer solution comprises the salts of between about 0.02M to about 0.04 M, the a self-assembled peptide _(IEIK) 3 I (SEQ ID NO: 2) The kit of claim 101.
(Item 104)
101. The kit of item 101, wherein said buffer comprises between about 0.1 M to about 0.4 M salt, and wherein said self-assembling peptide is (KLDL) ₃ (SEQ ID NO: 3).
(Item 105)
73. The kit according to item 72, wherein the subject is a mammal.
(Item 106)
106. The kit according to item 105, wherein the subject is a human.
(Item 107)
73. The kit according to item 72, wherein the self-assembling peptide comprises about 12 to about 16 amino acids in which hydrophobic amino acids and hydrophilic amino acids alternate.
(Item 108)
73. The kit according to item 72, further comprising at least one biologically active agent.
(Item 109)
73. The kit according to item 72, wherein the solution is substantially free of cells and drugs.
(Item 110)
73. The kit according to item 72, further comprising a sucrose solution.
(Item 111)
A composition comprising a self-assembling peptide comprising an effective amount and effective concentration of about 7 to 32 amino acids for use in forming a hydrogel barrier under physiological conditions to treat lung leakage.
(Item 112)
The hydrogel barrier, resulting in rupture pressure resistant of at least 35H ₂ O, The composition of claim 111.
(Item 113)
111. The self-assembling peptide according to item 111, wherein the self-assembling peptide is selected from the group consisting of (RADA) ₄ (SEQ ID NO: 1), (IEIK) ₃ I (SEQ ID NO: 2), and (KLDL) ₃ (SEQ ID NO: 3). Composition.
(Item 114)
Item 113, wherein the concentration effective to enable treatment of the lung leak comprises a self-assembling peptide concentration ranging from about 0.1 weight / volume (w / v) percent to about 3 w / v percent. A composition as described.
(Item 115)
111. The composition according to item 111, wherein the solution is substantially free of cells.
(Item 116)
111. The composition according to item 111, wherein the solution is substantially free of drug.
(Item 117)
112, further comprising at least one cation selected from the group consisting of ammonium ion, iron ion, magnesium ion, potassium ion, pyridinium ion, quaternary ammonium ion, sodium ion, potassium ion, and calcium ion. Composition.
(Item 118)
Selected from the group consisting of chloride, sulfate, acetate, carbonate, chloride, citrate, cyanide, fluoride, sulfate, nitrate, nitrite, and phosphate ions 111. The composition according to item 111, further comprising at least one anion.
(Item 119)
118. The composition according to item 117, comprising at least one of calcium chloride, sodium chloride, and potassium chloride.
(Item 120)
115. The composition according to item 114, comprising (RADA) ₄ (SEQ ID NO: 1) at a concentration of about 0.5 weight / volume (w / v) percent.
(Item 121)
121. The composition according to item 120, comprising a calcium chloride concentration of about 0.125M.
(Item 122)
122. The composition according to item 121, having a storage modulus of about 25 Pa.
(Item 123)
121. The composition according to item 120, comprising a calcium chloride concentration of about 0.250M.
(Item 124)
123. The composition according to item 123, having a storage modulus of about 44 Pa.
(Item 125)
121. The composition according to item 120, comprising a calcium chloride concentration of about 0.500M.
(Item 126)
125. The composition according to item 125, having a storage modulus of about 52 Pa.
(Item 127)
115. The composition of item 114 comprising (RADA) ₄ (SEQ ID NO: 1) at a concentration of about 2.5 weight / volume (w / v) percent.
(Item 128)
128. The composition according to item 127, comprising a calcium chloride concentration of about 0.125M.
(Item 129)
128. The composition according to item 128, having a storage modulus of about 600 Pa.
(Item 130)
115. The composition according to item 114, comprising a salt concentration between about 0.005M and about 1M.
(Item 131)
130. The composition according to item 130, comprising a salt concentration between about 0.125M and about 0.500M. (Item 132)
131. The composition according to item 131, comprising a salt concentration of about 0.25M.
(Item 133)
115. The composition according to item 114, further comprising a solution comprising sodium chloride, potassium chloride, calcium chloride, and sodium bicarbonate.
(Item 134)
115. The composition according to item 114, further comprising a solution comprising a contrast agent.
(Item 135)
135. The composition according to item 134, wherein said contrast agent comprises sulfate ions and sodium ions.
(Item 136)
111. The composition according to item 111, wherein the composition has a pH of about 2.5 to about 4.0.
(Item 137)
136. The method according to item 136, wherein the solution has a pH of about 3.5 and the self-assembling peptide is one of (RADA) ₄ (SEQ ID NO: 1) and (KLDL) ₃ (SEQ ID NO: 3). Composition.
(Item 138)
136. The composition according to item 136, wherein the solution has a pH of about 3.7 and said self-assembling peptide is (IEIK) ₃ I (SEQ ID NO: 2).
(Item 139)
111. The composition according to item 111, further comprising a buffer.
(Item 140)
140. The composition according to item 139, wherein the buffer comprises at least two salts.
(Item 141)
140. The composition according to item 139, wherein the buffer has a pH of 7.2.
(Item 142)
140. The composition according to item 139, wherein the buffer has a pH of 7.4.
(Item 143)
140. The composition according to item 139, wherein the buffer is an alkaline buffer.
(Item 144)
111. The composition according to item 111, comprising at least one of sodium chloride, potassium chloride, magnesium chloride, and calcium chloride at about 0.15M.
(Item 145)
144. The composition according to item 144, comprising between about 0.6M to about 1.2M of the salt, wherein the self-assembling peptide is (RADA) ₄ (SEQ ID NO: 1).
(Item 146)
As salt of between about 0.02M to about 0.04 M, the self-assembling peptide is a _(IEIK) 3 I (SEQ ID NO: 2) The composition of claim 144.
(Item 147)
144. The composition according to item 144, comprising between about 0.1 M to about 0.4 M salt, wherein the self-assembling peptide is (KLDL) ₃ (SEQ ID NO: 3).
(Item 148)
111. The composition according to item 111 for use in treating pulmonary leakage in a subject.
(Item 149)
111. The composition according to item 111, wherein the subject is a mammal.
(Item 150)
150. The composition according to item 149, wherein the subject is a human.
(Item 151)
111. The composition of item 111, wherein said self-assembling peptide comprises from about 12 to about 16 amino acids, wherein hydrophobic amino acids and hydrophilic amino acids alternate.
(Item 152)
111. The composition according to item 111, further comprising at least one biologically active agent.
(Item 153)
A method of facilitating treatment of lung leakage in a subject, comprising:
Providing a solution comprising an effective amount and concentration of a self-assembling peptide comprising from about 7 amino acids to about 32 amino acids to form a hydrogel barrier under physiological conditions to enable treatment of said lung leakage Steps and
Providing instructions for administering the solution to a target area of the lung leak by introducing the solution through a delivery device located at the leak of the lung.
(Item 154)
153. The method of item 153, further comprising providing an instruction to visualize an area that includes at least a portion of the lung leak.
(Item 155)
Providing instructions for visualizing an area comprising at least a portion of the lung leakage,
Identifying the target area of leakage of the lung;
Introducing the delivery device;
Placing the distal end of the delivery device within the target area;
Administering the solution;
Removing the delivery device from the lung leak; and monitoring the lung leak after removing the delivery device;
154. The method of claim 154, comprising providing an indication to visualize the region during at least one of the following.
(Item 156)
157. The method of item 154, further comprising providing an instruction to visualize the area within a time period of about 1 minute to about 5 minutes after administering the solution.
(Item 157)
153. The method of item 153, further comprising providing instructions for preparing the effective amount and / or the effective concentration based in part on a size of the target area of the lung leak.
(Item 158)
157. The method according to item 157, wherein the effective amount is approximately 1 mL / cm ^{2 of} target area.
(Item 159)
153. The self-assembling peptide according to item 153, wherein the self-assembling peptide is selected from the group consisting of (RADA) ₄ (SEQ ID NO: 1), (IEIK) ₃ I (SEQ ID NO: 2), and (KLDL) ₃ (SEQ ID NO: 3). the method of.
(Item 160)
160. The method of item 159, wherein the concentration effective to allow prevention of lung leakage comprises a concentration in the range of about 0.1% w / v to about 3% w / v peptide.
(Item 161)
170. The method of item 160, wherein the amount effective to allow prevention of lung leakage comprises a volume in the range of about 0.1 mL to about 5 mL.
(Item 162)
153. The method of item 153, further comprising providing instructions for monitoring an area surrounding the target area.
(Item 163)
153. The method of item 153, further comprising providing the solution and instructions for use after a surgical procedure.
(Item 164)
Providing a solution comprising a self-assembling peptide comprises instructions for preparing a peptide solution having an effective concentration to form a hydrogel barrier under physiological conditions to allow for prevention of lung leakage. 153. The method of item 153, comprising providing
(Item 165)
A macroscopic scaffold consisting essentially of a plurality of self-assembled peptides, wherein each of said self-assembled peptides has an effective amount of from about 7 amino acids to about 7 amino acids capable of being located within a target area of lung leakage. A macroscopic scaffold containing 32 amino acids.
(Item 166)
165, wherein each of said plurality of peptides comprises one of (RADA) ₄ (SEQ ID NO: 1), (IEIK) ₃ I (SEQ ID NO: 2), and (KLDL) ₃ (SEQ ID NO: 3). Scaffold visible to the naked eye as described.
(Item 167)
167. A macroscopic scaffold according to item 166, comprising nanofibers having a diameter from about 10 nanometers to about 20 nanometers.

  Kits are provided for treating lung leakage in a subject. The kit includes a self-assembling peptide comprising an effective amount of about 7 amino acids to about 32 amino acids to form a hydrogel barrier under physiological conditions to enable treatment of lung leakage. The kit also includes instructions for administering the self-assembling peptide to the target area of the lung leak in the subject.

  Provided is a composition comprising a self-assembling peptide comprising an effective amount and effective concentration of about 7 to 32 amino acids for use in forming a hydrogel barrier under physiological conditions to treat lung leakage. Is done. In embodiments, methods are provided that facilitate preventing lung leakage in a subject. The method provides a solution comprising an effective amount and an effective concentration of a self-assembling peptide comprising from about 7 amino acids to about 32 amino acids to form a hydrogel barrier under physiological conditions that allows for prevention of lung leakage. Including the step of: The method also includes the step of providing instructions for administering the solution to the target area of the lung leak by introducing the solution through a delivery device disposed within the leak of the lung.

  A macroscopic scaffold consisting essentially of a plurality of self-assembling peptides is provided. Each of the self-assembling peptides contains an effective amount of about 7 amino acids to about 32 amino acids that can be located within the target area of lung leakage.

1A and 1B are graphs showing data considered in Example 1. FIG. 2 to 4 are graphs showing data considered in Example 3. 2 to 4 are graphs showing data considered in Example 3. 2 to 4 are graphs showing data considered in Example 3. 5A and 5B are graphs showing data considered in Example 3. 6A-6B are graphs showing data considered in Example 4. 7A and 7B are graphs showing data considered in Example 5. 8A-8B are graphs showing data considered in Example 5. FIG. 9A-9B are graphs showing data considered in Example 6. 10 to 12 are graphs showing data considered in Example 8. 10 to 12 are graphs showing data considered in Example 8. 10 to 12 are graphs showing data considered in Example 8. 13 and 14 are graphs showing data considered in Example 9. 13 and 14 are graphs showing data considered in Example 9. 15 and 16 are graphs showing the data considered in Example 10. 15 and 16 are graphs showing the data considered in Example 10. 17 and 18 are graphs showing data considered in Example 11. 17 and 18 are graphs showing data considered in Example 11. FIG. 19 is a graph showing data considered in Example 12. FIG. 20 is a graph showing the data considered in Example 13. FIG. 21 is a graph showing data considered in Example 14. FIG. 22 is a graph showing data considered in Example 15. 23A to 23B are graphs showing data considered in Example 16. 24A to 24C are graphs showing data considered in Example 18. 24A to 24C are graphs showing data considered in Example 18. FIG. 25 is a diagram illustrating data considered in Example 19. FIG. 25 discloses “IEIK” of SEQ ID NO: 5. FIG. 26 is a graph showing data considered in Example 19. FIG. 27 is a graph showing data considered in Example 19. FIG. 28 is a graph showing the data considered in Example 19. FIG. 29 is a graph showing data considered in Example 19. 30A to 30C are graphs showing data considered in Example 19. FIG. 30 discloses “IEIK” of SEQ ID NO: 5.

  The systems and methods of the present disclosure can facilitate prevention of pulmonary leaks, for example, pleural defects or bronchial anastomotic leaks.

  Pleural defects, such as parenchymal parenchymal defects, are often created by partial excision, synectomy, or interlobar transection. In up to 25% of patients, persistent postoperative air leaks for more than 7 days have been reported. Controlling these air leaks during surgery reduces morbidity and length of hospital stay.

  Leakage of the bronchial anastomosis is generally observed post-operatively during the examination. Leakage of the bronchial anastomosis can cause severe complications and may require additional surgery or procedures. Conventionally, the anastomotic site is sutured directly to prevent leakage of the bronchial anastomosis, and then dabbed using an omental flap, an intercostal muscle flap, or a pericardial fat flap. Do that. Currently, no material has been approved for clinical use to prevent bronchial anastomosis leakage.

  According to one or more embodiments, the self-assembling peptide can treat a leak in the lung, for example, a pleural defect or bronchial anastomosis. Leakage can be created after surgery. In some non-limiting embodiments, the materials, systems and methods disclosed herein can facilitate mucosal epithelialization.

  Self-assembling peptides of the present disclosure may include applications, eg, administration of the self-assembling peptide to a predetermined or desired target area. The self-assembling peptide can be administered to the target area in the form of a peptide solution, hydrogel, membrane or other form. The target area may be a predetermined area of the subject in need of a particular treatment. In some embodiments, the target area may be associated with a surgical site (eg, a sleep-breathing disorder (SBD) surgical site).

  During self-assembly, the peptides can form nanofibers. Self-assembly can cause gelation of the peptide in solution. Gelation can result in or form a hydrogel. Peptides can spontaneously form beta-sheets at neutral pH levels in solution. Peptides may spontaneously form beta-sheets in solution, under physiological conditions and / or in the presence of cations and / or anions.

  The methods and materials of the present disclosure can be used after a surgical procedure. For example, a solution containing a self-assembling peptide can be administered after a surgical procedure.

  The methods of the present disclosure may include introducing a delivery device to a target area of a leak in a subject's lungs. The method can include placing the distal end of the delivery device within a target area where treatment of a lung leak is desired.

  The methods of the present disclosure may also include administering the self-assembling peptide to a predetermined or desired target. The self-assembling peptide can be administered to the target area in the form of a peptide solution, hydrogel, membrane or other form. The target area may be a predetermined area of the subject in need of a particular treatment. In some embodiments, the target area may relate to a surgical site or a site of lung leakage. Leakage of the lungs may be the result of a pleural defect or of a bronchial anastomosis. For example, the surgical site may be a site where a surgical operation such as a lung-related surgery has been performed, or a site where a pleural defect or bronchial anastomosis leak has occurred. The pleural defect may be associated with a needle puncture, a staple line, or a suture line. A pleural defect can be less than about 5 x 5 mm.

  Administration to the target area through the delivery device can be performed to form a hydrogel barrier. This can be done under physiological conditions in the target area to treat lung leakage.

  Materials and methods can include treating, preventing, or occluding a leak in the lungs.

  As used herein, the term "treatment" is intended to include a leak, for example, a partial or complete occlusion or blockage of an area where an air leak has occurred. In general, leakage is undesirable, and the treatment corrects the leak and results in healing of the targeted area of treatment. Treatment of a subject can include one or more of healing, alleviating, alleviating or improving a subject having a disorder, eg, leakage, than would be expected in the absence of such treatment. The treatment may be a minimally invasive treatment, including minimally invasive application or administration of a solution containing the self-assembling peptide.

  As used herein, the term "subject" is intended to include human and non-human animals, such as vertebrates, large animals, and primates. In certain embodiments, the subject is a mammalian subject, and in certain embodiments, the subject is a human subject. While applications in humans are clearly anticipated, veterinary applications using, for example, non-human animals are also contemplated in the present invention. The term “non-human animal” of the present invention includes all vertebrates, eg, non-mammals (eg, birds, eg, chickens; amphibians; reptiles, etc.) and mammals, eg, non-human primates, livestock, And agriculturally useful animals such as sheep, dogs, cats, cows, pigs, rats and the like.

  The treatment, prevention or occlusion may be partial or complete. Materials and methods can include, for example, addressing pulmonary leaks from pleural defects or bronchial anastomotic leaks. Materials and methods can include administering, applying, or injecting a self-assembling peptide, or a solution comprising the self-assembling peptide, or a composition comprising the self-assembling peptide, to a predetermined or desired target area.

  The method of treating a lung leak can further include removing the delivery device from the target area. The method may further include, before the step of introducing the delivery device, visualizing an area including the target area. Visualization of the area containing the target area can be performed after the step of removing the delivery device from the target area. The step of monitoring the target area can also be performed during and after the procedure, for example, after removing the delivery device.

  The method of treatment may further comprise the step of preparing a solution comprising the self-assembling peptide. In some embodiments, the method of treatment can further include assessing the subject to determine the need to treat a lung leak and preparing a solution based on the assessing.

  The term “self-assembling peptide” can refer to a peptide that can exhibit a beta-sheet structure in aqueous solution in the presence of certain conditions under which the beta-sheet structure is induced. These particular conditions may include adjusting the pH of the self-assembling peptide solution. Adjustment may be to raise or lower the pH of the self-assembling peptide solution. Raising the pH may be raising the pH to a physiological pH. Certain conditions may also include adding a cation, such as a monovalent cation or a divalent cation, to the self-assembling peptide solution. Certain conditions may also include adding an anion, such as a monovalent or divalent anion, to the self-assembling peptide solution. Particular conditions may include those associated with the site of surgery or the target site of lung leakage. The self-assembling peptide may be referred to as, or be part of, a composition, peptide solution, peptide powder, hydrogel, or scaffold.

  The term “self-assembling peptide” can refer to a peptide that contains a self-assembling motif. Self-assembling peptides are peptides that can self-assemble into structures, including but not limited to macroscopic membranes or nanostructures.

  The term "hydrogel" may refer to a material composed of a polymer and a high percentage of water, for example, at least 90% water.

  The self-assembling peptide may be an amphiphilic self-assembling peptide. "Amphiphilic" means that the peptide contains a hydrophobic moiety and a hydrophilic moiety. In some embodiments, the amphipathic peptide may comprise, consist essentially of, or consist of alternating hydrophobic and hydrophilic amino acids. The term "alternating" includes a series of three or more amino acids in which hydrophobic amino acids and hydrophilic amino acids alternate, and each of the peptide sequences in which hydrophobic amino acids and hydrophilic amino acids alternate appears. It is not necessary to include all amino acids. Self-assembling peptides, also referred to herein as "peptides", are administered to a predetermined or desired target area in the form of a self-assembling peptide solution, composition, hydrogel, membrane, scaffold or other form. be able to. Hydrogels can also be referred to as membranes or scaffolds throughout this disclosure. The predetermined or desired target area may be at or near a lung leak (eg, a pleural defect or a bronchial anastomotic leak). The predetermined or desired target area may be established based on a surgical procedure or a site or other area that may have been unintentionally or intentionally traumatized.

  The solution containing the self-assembling peptide is also called a self-assembling peptide solution, and may be an aqueous self-assembling peptide solution. The self-assembling peptide can be administered, applied, or injected in a substantially cell-free or cell-free solution. In certain embodiments, the self-assembling peptide can be administered, applied, or injected in a cell-free or cell-free solution.

  The self-assembling peptide can also be administered, applied, or injected in a substantially drug-free or drug-free solution. In certain embodiments, the self-assembling peptide can be administered, applied, or injected in a drug-free or drug-free solution. In certain other embodiments, the self-assembling peptide can be administered, applied, or injected in a substantially cell-free, substantially drug-free solution. In certain further other embodiments, the self-assembling peptide can be administered, applied, or injected in a cell-free, drug-free solution.

  The self-assembling peptide solution may comprise, consist of, or consist essentially of the self-assembling peptide. The self-assembling peptide may be in a modified or unmodified form. Modified means that the self-assembling peptide can have one or more domains that include one or more amino acids that do not self-assemble when provided alone in solution. Unmodified means that the self-assembling peptide may not have any other domains other than those that effect the self-assembly of the peptide. That is, the unmodified peptide consists of alternating hydrophobic and hydrophilic amino acids that can self-assemble into macroscopic structures such as beta-sheets and hydrogels.

  By administering the solution containing the self-assembling peptide, a hydrogel barrier can be formed. A hydrogel barrier can be formed within the target area to treat lung leakage. Treatment can be effected by at least partially occluding or sealing the lung leak. This is achieved by forming a hydrogel barrier. Throughout this disclosure, reference to a hydrogel may refer to or be applicable to a hydrogel barrier.

  In certain embodiments, it is desirable to have a hydrogel barrier that can provide a suitable or desired blockage or seal at the target area. The hydrogel barrier may have certain properties to achieve a suitable or desired barrier or seal. For example, a hydrogel barrier has one or more predetermined properties, such as mechanical strength (storage modulus), stiffness, viscosity, gelation kinetics, ionic strength, pH, or burst pressure (burst pressure resistance). May be something. The properties can be adjusted or adapted based on the addition of the components disclosed herein in specific amounts and / or concentrations to the self-assembling peptide or a solution containing the self-assembling peptide. it can.

In certain embodiments, treated by at least 20 cmH ₂ O, at least 25 cmH ₂ O, at least 30 cm H ₂ O, and in certain instances can result in the least 35cmH ₂ O burst pressure.

  For example, for treating lung leakage, it is desirable to provide a hydrogel barrier with high mechanical strength, stiffness, and high burst pressure. It is also desirable to provide a hydrogel barrier that gels rapidly, that is, the gelation kinetics is such that once administered, the hydrogel barrier forms within a short period of time and the leak is treated. A short time can be immediate, such as less than 5 minutes, less than 3 minutes, less than 2 minutes, less than 1 minute, or less than 30 seconds, or any other time disclosed herein. Is also good.

  The administering of the solution may comprise, consisting of, consisting of, a solution comprising, consisting of, or consisting essentially of a self-assembling peptide comprising, consisting of, or consisting essentially of at least about 7 amino acids, Or it may be essentially from it. The administration of the solution may comprise the administration of a solution comprising, consisting of, or consisting essentially of a self-assembling peptide comprising, consisting of, or consisting essentially of about 7 amino acids to 32 amino acids, consisting of Good or may be essentially from it. Other peptides that do not comprise, consist of, or consist essentially of at least about 7 amino acids may also be contemplated in the present disclosure.

  A self-assembling peptide may comprise, consist of, or consist essentially of about 7 to about 32 amino acids. In some embodiments, a self-assembling peptide may comprise, consist of, or consist essentially of about 12 to about 16 amino acids.

  The term "alternating" includes a series of three or more amino acids in which hydrophobic amino acids and hydrophilic amino acids alternate, and each of the peptide sequences in which hydrophobic amino acids and hydrophilic amino acids alternate appears. It is not necessary to include all amino acids.

  A method for treating lung leakage may include administering a self-assembling peptide to a target area. The peptide may be administered as a hydrogel, or may form a hydrogel upon administration. A method of treating pulmonary leakage may include administering a solution comprising a self-assembling peptide to a target area.

  The term `` administering '' refers to a variety of forms, including, but not limited to, self-assembling peptides, alone, as a solution, such as an aqueous solution, or as a composition, hydrogel, or scaffold. It shall include applying, introducing or injecting in one or more of the forms, with or without additional components.

  The method may include introducing a delivery device to a target area of the lung leak in the subject. The method may include introducing a delivery device including at least one of a syringe, tube, pipette, catheter, catheter syringe, or other needle-based device to a target area of the subject. The self-assembling peptide can be administered to a subject's target area by a syringe, tube, pipette, catheter, catheter syringe, or other needle-based device. The gauge of the syringe needle can be selected to provide an appropriate flow of the composition, solution, hydrogel, or liquid from the syringe to the target area. This means, in some embodiments, that the amount of the self-assembling peptide in the composition, peptide solution, or hydrogel to be administered, the concentration of the peptide solution in the composition or hydrogel, and the peptide solution, the composition Or based on at least one of the viscosities of the hydrogels. The delivery device can be a conventional device that can reach a particular target area, achieve a particular dosing regimen, deliver a particular target volume, amount, or concentration, and deliver a precise target area. May be designed to achieve at least one of the following:

  A method of treating a lung leak may include placing the distal end of the delivery device within a target area where treatment of the lung leak is desired. The target area may be an area described herein, such as a portion of a surgical site, a site of a pleural defect or a site of bronchial anastomosis leak. The self-assembling peptide can be administered to the target area where treatment of the leak is desired by the delivery device. The self-assembling peptide can be administered to the target area by a delivery device in solution. In some embodiments, the administration is performed locally, in that the delivery device is located in close proximity to the target area or leak, resulting in a solution containing the self-assembling peptide at the surface of the target area or at the location of the leak. be able to. In other embodiments, administration can be directly to the leak, in that the delivery device is placed at the target area or leak and the solution containing the self-assembling peptide is brought directly to the target area or location of the leak, for example. it can. In other embodiments, administration can be to a target area or leak into or through a leak to fill a predetermined target area volume with a solution comprising a self-assembling peptide. Administering the solution may include applying the solution locally to the target area. Administering the solution may include injecting the solution into the target area by overflow to locally cover the target area.

  The use of a delivery device can result in more selective administration of the peptide, resulting in more accurate delivery to the target area. Selective administration of the peptide may allow for an enhanced and more targeted delivery of the peptide solution, composition, or hydrogel, which is successful and precisely located at the desired location. Selective administration can result in enhanced, targeted delivery with significantly improved placement and therapeutic effects over the use of alternative delivery devices. Delivery devices that can be used in the systems, methods, and kits of the present disclosure include syringes, tubes, needles, pipettes, syringe catheters, other needle-based devices, or catheters.

  The use of a delivery device, such as a catheter, allows for the proper placement of a guidewire or catheter or other device used to guide the catheter to a location, and visualization of the target area and / or path to the target area. It may include the use of ancillary devices such as endoscopes. The endoscope may be a tube that may include at least one of light and a camera or other visualization device to allow viewing of an image of the subject's body. A guidewire or endoscope can be introduced into a subject by, for example, incising the skin. The endoscope can be introduced to the target area prior to introducing the delivery device to the target area.

  Use of delivery devices such as syringes, tubes, needles, pipettes, syringe catheters, other needle-based devices, catheters, or endoscopes include syringes, tubes, needles, pipettes, syringe catheters, other needle-type devices, The diameter of the opening of the catheter or endoscope is such that at least a portion of the endoscope is penetrated into the opening where the target area is present and the peptide, peptide solution, composition or hydrogel can be administered to the target area. It may be necessary to determine the size.

  In certain embodiments, the hydrogel can be formed in vitro and administered in vivo to a desired location. In certain instances, this location may be a target area. In other examples, the location may be upstream, downstream, or substantially near the area. It is desirable to allow the transfer of the hydrogel to the area where it is desired to move. Alternatively, the hydrogel can be placed in the desired area by another procedure. The desired location or target area can be at least a portion of an area where it is desired to treat a lung leak in a subject.

  In certain aspects of the present disclosure, the hydrogel can be formed in vivo. To treat lung leakage in a subject, a solution containing a self-assembling peptide, such as an aqueous solution, can be inserted into the subject's in vivo location or area. In certain instances, the hydrogel is formed in one location in vivo and treated for pulmonary leakage in a subject, eg, blockage or obstruction at or near lung leakage (eg, pleural deficiency or bronchial anastomosis). Can be moved to an area where it is desired to promote or bring about. Alternatively, another procedure may place the hydrogel in an area where it is desired to facilitate or effect the treatment of a lung leak. The peptides of the present disclosure may be in the form of a powder, a solution, a gel, and the like. The self-assembling peptide gels in response to changes in the pH and salt concentration of the solution and can be distributed as a liquid that gels upon contact with the subject during application or administration.

  In certain circumstances, the peptide solution may be a weak hydrogel, and consequently can be administered by the delivery devices described herein.

  According to some embodiments, the self-assembling peptide can be amphiphilic, where hydrophobic amino acids and hydrophilic amino acids alternate.

  According to one or more embodiments, the subject can be evaluated to determine the need to treat a lung leak in the subject. Upon completion of the evaluation, a peptide solution to be administered to the subject can be prepared based on the step of evaluating. In other embodiments, peptide solutions can be prepared without the step of evaluating.

  In some embodiments, biologically active agents can be used with the materials and methods of the present disclosure. Biologically active agents are peptides, DNA sequences, chemical compounds, or inorganic or organic compounds that can provide some activity, control, regulation, or modulation of conditions or other activities in a subject or laboratory setting. May be included. A biologically active agent can interact with another component to provide such activity. A biologically active agent can be referred to herein as a drug, according to some embodiments. In certain embodiments, one or more biologically active agents can be slowly released outside the peptide system. For example, one or more biologically active agents can be slowly released from the hydrogel. Both in vitro and in vivo studies have demonstrated this gradual release of biologically active agents. The biologically active agent can also be added to the self-assembling peptide solution or composition prior to administration to the subject, and can be added to the subject in conjunction with the self-assembling peptide or separately from the self-assembling peptide. It can also be administered. One or more biologically active agents can be encapsulated in the system, for example, in a hydrogel, solution, composition, or nanofiber.

  The present disclosure relates to aqueous solutions, hydrogels, scaffolds, compositions and membranes containing self-assembling peptides, sometimes referred to as self-assembling oligopeptides. The self-assembling peptide is applied in aqueous solution, at physiological pH and / or cations and / or anions, such as monovalent cations and / or monovalent anions, or at or near the site of surgery or leaks in the lung. Beta-sheet structures may be exhibited in the presence of other possible conditions. The peptide may be an amphiphilic, hydrophobic and hydrophilic amino acid alternating. In certain embodiments, the peptide may include a first portion that may be amphipathic, where hydrophobic and hydrophilic amino acids alternate, and another portion or region that is not amphipathic.

  Peptides are generally stable in aqueous solution and may self-assemble into large, macroscopic structures, scaffolds, or matrices when exposed to selected conditions. The conditions can be physiological conditions, neutral pH, selected salt concentrations, buffered solutions, or physiological levels of salt. Once the hydrogel is formed, the hydrogel may not be degraded and may degrade or biodegrade after a period of time. The rate of degradation may be based, at least in part, on at least one of the amino acid sequence and surrounding conditions. The rate of degradation can be correlated with the rate of healing or growth at the target site to provide adequate treatment of lung leakage.

  By "visible to the naked eye" is meant having sufficiently large dimensions that they can be seen under a magnification of 10 or less. In a preferred embodiment, the macroscopic structure is visible to the naked eye. The structure visible to the naked eye may be transparent and may be two-dimensional or three-dimensional. Generally, each dimension is at least 10 μm in size. In certain embodiments, at least two dimensions are at least 100 μm, or at least 1000 μm in size. Often, the at least two dimensions are at least 1-10 mm in size, 10-100 mm in size, or more.

  In certain embodiments, the size of the filament may be from about 10 nanometers (nm) to about 20 nm. The distance between the filaments can be from about 50 nm to about 80 nm.

The macroscopic structure may be a macroscopic scaffold. The macroscopic scaffold may consist essentially of a plurality of self-assembling peptides. Each of the self-assembling peptides may comprise an effective amount of about 7 amino acids to about 32 amino acids that can be placed in the target area of the pulmonary system to prevent pulmonary leakage, and then comprises Or may consist of it. The self-assembling peptide of the scaffold may comprise from about 12 to about 16 amino acids. The self-assembling peptide of the scaffold may comprise from about 12 to about 16 amino acids alternating between hydrophobic and hydrophilic amino acids. The self-assembled peptide may comprise (RADA) ₄ (SEQ ID NO: 1), (IEIK) ₃ I (SEQ ID NO: 2), (KLDL) ₃ (SEQ ID NO: 3), and may consist essentially of: It may consist of it.

  "Physiological conditions" can occur naturally with respect to a particular organism, cell line, or subject, and can be in contrast to artificial laboratory conditions. Conditions may include one or more properties, such as one or more particular properties or a range of one or more properties. For example, physiological conditions may include temperature or a range of temperatures, pH or a range of pHs, pressures or ranges of pressures, and concentrations of one or more of certain compounds, salts, and other components. The salt may include one or more of a monovalent anion, a monovalent cation, a divalent anion, or a monovalent cation.

  In some examples, the physiological condition may include a temperature in a range from about 20 to about 40 degrees Celsius. In some examples, the pressure may be about 1 atm. The pH may be in the range of a neutral pH. For example, the pH may range from about 6 to about 8. Physiological conditions may include cations and / or anions such as monovalent metal cations and / or monovalent anions that can induce the formation of a membrane or hydrogel. These may include sodium chloride (NaCl). Physiological conditions may also include glucose, sucrose, or other sugar concentrations between about 1 mM and about 20 mM. The self-assembling peptide solution may include glucose, sucrose, or other sugar, and the sugar or sugar solution may be added to the self-assembling peptide solution.

  In certain embodiments, a self-assembling peptide may be a peptide of at least about 7 amino acids. In certain other embodiments, the self-assembling peptide can be a peptide of at least about 7 amino acids to about 32 amino acids. In certain other embodiments, the self-assembling peptide may be a peptide of about 7 to about 17 amino acids. In certain other examples, the self-assembling peptide may be a peptide of at least 8 amino acids, at least about 12 amino acids, or at least about 16 amino acids.

  Both homogeneous and heterogeneous mixtures of peptides characterized by the above properties can form stable macroscopic membranes, filaments, and hydrogels. Self-complementary, self-compatible peptides can form membranes, filaments, and hydrogels in a homogeneous mixture. Heterogeneous peptides, including those that cannot form membranes, filaments, and hydrogels in homogenous solutions that are complementary to each other and / or are structurally compatible, also include macroscopic membranes, Can self-assemble into filaments and hydrogels.

  The membranes, filaments, and hydrogels can be non-cytotoxic. The hydrogels of the present disclosure may be digested and metabolized in a subject. The hydrogel may be biodegradable in 30 days or less. Hydrogels have a simple composition, are permeable, are easy to make in large quantities, and are relatively inexpensive. The membrane and filament, hydrogel or scaffold can also be made and stored under sterile conditions. The optimal length for film formation can vary with at least one of the amino acid composition, solution conditions, and conditions in the target area.

  The amino acids of the self-assembled or amphiphilic peptide can be selected from D-amino acids, L-amino acids, or combinations thereof. Hydrophobic amino acids include Ala, Val, Ile, Met, Phe, Tyr, Trp, Ser, Thr and Gly. The hydrophilic amino acid may be a basic amino acid, for example, Lys, Arg, His, Orn; an acidic amino acid, for example, Glu, Asp; or an amino acid that forms a hydrogen bond, for example, Asn, Gln. Acidic and basic amino acids can be clustered on the peptide. The carboxyl and amino groups of the terminal residue may be protected or unprotected. Membranes or hydrogels can be formed in a homogeneous mixture of self-complementary and self-compatible peptides or in a heterogeneous mixture of peptides that are complementary and structurally compatible with each other. Peptides that meet the above criteria may self-assemble into a macroscopic membrane under the appropriate conditions described herein.

  In certain embodiments, about 8 to about 32 residues can be used for a self-assembling peptide, and in other embodiments, the self-assembling peptide can have about 7 to about 17 residues. . The length of the peptide may be about 5 nm.

  The peptides of the present disclosure may comprise, or consist essentially of, peptides having the arginine, alanine, aspartic acid and alanine repeats (Arg-Ala-Asp-Ala (RADA) (SEQ ID NO: 4)). May consist of it.

Another peptide sequence is a self-assembling peptide comprising, consisting essentially of, or consisting of a repetitive sequence of isoleucine, glutamic acid, isoleucine, and lysine (Ile-Glu-Ile-Lys (IEIK) (SEQ ID NO: 5)). Can be represented. Other peptide sequences include, consist essentially of, or consist of self-assembly of lysine, leucine, aspartic acid, and the repetitive sequence of leucine (Lys-Leu-Asp-Leu (KLDL) (SEQ ID NO: 6)). It can be represented by a peptide. As a specific example of a self-assembling peptide according to the present invention, the sequence Arg-Ala-Asp-Ala-Arg-Ala-Asp-Ala-Arg-Ala-Asp-Ala-Arg-Ala-Asp-Ala (SEQ ID NO: 1) A self-assembling peptide referred to as “RADA16” with ((RADA) ₄ (SEQ ID NO: 1)) (also referred to as “Puramatrix” throughout this disclosure), the sequence Ile-Glu-Ile-Lys-Ile-Glu. -Ile-Lys-Ile-Glu- Ile-Lys-Ile ( SEQ ID NO: _{2) ((IEIK) 3 I} ( SEQ ID NO: 2)) self-assembling peptides called "IEIK13" having or sequence Lys-Leu, -Asp-Leu-Lys-Leu-Asp-Leu-Lys-Leu-Asp-Leu (SEQ ID NO: 3) ( There can be a self-assembling peptide referred to as “KLDL12” having (KLDL) ₃ (SEQ ID NO: 3)) (which can also be referred to as “KLD12” throughout this disclosure).

  Each of the peptide sequences disclosed herein can provide a peptide comprising, consisting essentially of, and consisting of the recited amino acid sequence.

  The present disclosure provides materials, methods, and kits for solutions, hydrogels, compositions, and scaffolds comprising, consisting essentially of, or consisting of the peptides listed herein.

One weight / volume (w / v) percent aqueous (water) solution and 2.5 w / v percent (RADA) ₄ (SEQ ID NO: 1) were obtained from 3-D Matrix Co. , Ltd., available as a product PuraMatrix ™ peptide hydrogel.

  The self-assembly of the peptide may be caused by hydrogen bonds and hydrophobic bonds between peptide molecules by amino acids constituting the peptide.

  The diameter of the self-assembling peptide nanofibers of the present disclosure ranges from about 10 nm to about 20 nm, and the average pore size ranges from about 5 nm to about 200 nm. In certain embodiments, the nanofiber diameter, pore size, and nanofiber density can be controlled by at least one of the concentration of the peptide solution used and the amount of the peptide solution used, such as the volume of the peptide solution. it can.

  As such, the specific concentration of the peptide in solution and the concentration to provide at least one of the desired nanofiber diameter, pore size, and density to properly effect treatment of lung leakage, e.g., to effect obstruction. At least one of a certain amount of the peptide solution can be selected. A particular concentration and a particular amount of a peptide solution can be referred to as an “effective concentration” and an “effective amount”.

  As used herein, an amount, "effective amount" or "therapeutically effective amount" of a peptide, peptide solution or hydrogel effective to treat pulmonary leakage in a subject refers to a subject having a disorder. In the treatment of, or in healing, alleviation, alleviation or improvement, when administered to a subject in a single dose or in multiple doses (application or injection), is more effective than would be expected in the absence of such treatment. Refers to the amount of a peptide, peptide solution, composition, or hydrogel. This may be a specific concentration or range of concentrations of the peptide in the peptide solution, composition or hydrogel, or additionally or alternatively, a specific volume or range of the volume of the peptide solution, composition or hydrogel. May be included. The method of facilitating can include providing instructions for preparing an effective amount and / or concentration.

  The dosage, eg, volume or concentration, to be administered (eg, applied or injected) depends on the form of the peptide (eg, peptide solution, hydrogel, or dry form, such as lyophilized form) and the route of administration utilized. Can fluctuate. The exact formulation, route of administration, volume, and concentration will be chosen in view of the subject's condition and the particular target area or location to which the peptide solution, hydrogel, or other form of peptide will be administered. be able to. Lower or higher doses than those listed herein may or need be used. The particular dosage and treatment regimen for any particular subject will result in the particular peptide (s) used, the size of the area to be treated, and the desired target area, which can be located within the desired target area. It can depend on a variety of factors, which can include the desired thickness of the hydrogel, and the length of treatment time. Other factors that may affect a particular dosage and treatment regimen include age, weight, overall health, gender, time of administration, rate of degradation, severity and course of the disease, condition or condition And the judgment of the treating physician. In certain embodiments, the peptide solution can be administered in a single dose. In other embodiments, the peptide solution can be administered in more than one dose, or in multiple doses.

  The effective amount and effective concentration of the peptide solution are selected so that lung leakage is at least partially treated, e.g., to promote or result in obstruction of a subject at or near the leak of a pleural defect or bronchial anastomosis. can do. In some embodiments, the effective amount and / or concentration may be based in part on the size or diameter of the target area. In other embodiments, the effective amount and / or concentration is based in part on the flow rate of one or more fluids at or near the target area. In yet other embodiments, the effective amount and / or concentration may be based in part on the size or diameter of the target area at the site of lung leakage or surgery.

  In yet other embodiments, the effective amount and / or concentration is at least one of the size or diameter of the target area, and the flow rate of one or more fluids at or near the target area, and leakage of the lung, such as a pleural defect or It may be based in part on at least one of the size or diameter of the target area at the site of the bronchial anastomosis leak or the surgical procedure.

  An effective amount can include a volume of the peptide solution from about 0.1 milliliter (mL) to about 100 mL. An effective amount can include from about 0.1 mL to about 10 mL of the peptide solution volume. An effective amount can include a volume from about 0.1 to about 5 mL. In certain embodiments, the effective amount can be about 0.4 mL. In certain embodiments, the effective amount can be about 0.5 mL. In other embodiments, the effective amount can be about 1.0 mL. In still other embodiments, the effective amount can be about 1.5 mL. In still other embodiments, the effective amount can be about 2.0 mL. In some other embodiments, the effective amount can be about 3.0 mL.

In certain embodiments, the effective amount may be a target area 1 cm ² per approximately 0.1ML～1cm ² per approximately 5 mL. An effective amount can be from about per 1cm ² 0.1mL~1cm ² per about 3 ml. In certain embodiments, the effective amount can be approximately 1 mL / cm ^{2 of} target area. This effective amount can be used in connection with a concentration such as 1.5% w / v or 2.5% w / v of the peptide solution of the present disclosure.

  An effective concentration can be an amount that can treat lung leakage, as described herein. The various properties at or near the target site may make the selection or determination of an effective concentration including at least one of the size or diameter of the target area and the flow rate of one or more fluids at or near the target area. Can be a factor.

  Effective concentrations can include peptide concentrations in the solution ranging from about 0.1 weight / volume (w / v) percent to about 10 w / v percent. Effective concentrations can include peptide concentrations in the solution ranging from about 0.1 w / v percent to about 3.5 w / v percent. In certain embodiments, the effective concentration can be about 1 w / v percent. In certain other embodiments, the effective concentration can be about 1.5 w / v percent. In another embodiment, the effective concentration can be about 2.5 w / v percent. In yet other embodiments, the effective concentration can be about 3.0 w / v percent.

  In certain embodiments, a peptide solution having a higher concentration of peptide can result in a more effective hydrogel that can stay in place and provide effective treatment. To deliver the peptide solution, a more concentrated peptide solution may become too viscous to allow for effective and selective solution administration. If a sufficiently high concentration is not selected, the hydrogel may not be able to have efficacy in the target area for the desired period of time.

  The effective concentration can be selected to provide a solution that can be administered by injection or other means using a catheter or needle of a particular diameter or gauge.

  The methods of the present disclosure contemplate single and multiple administrations of a therapeutically effective amount of the peptides, compositions, peptide solutions, membranes, filaments, and hydrogels described herein. The peptides described herein can be administered at regular intervals depending on the nature, severity and extent of the subject's condition. In some embodiments, the peptide, composition, peptide solution, membrane, filament, or hydrogel can be administered in a single dose. In some embodiments, the peptides, compositions, peptide solutions, or hydrogels described herein are administered in multiple doses. In some embodiments, a therapeutically effective amount of a peptide, composition, peptide solution, membrane, filament, or hydrogel can be administered periodically at regular intervals. The periodic interval selected may be based on any one or more of the initial peptide concentration of the administered solution, the amount administered, and the rate of degradation of the formed hydrogel. For example, after the first administration, the subsequent administration is performed, for example, 30 seconds, 1 minute, 2 minutes, 5 minutes, 10 minutes, 1 day, 2 days, 5 days, 1 week, 2 weeks This can be done after 4, 6 or 8 weeks. Subsequent administration may include administration of a solution in which the concentration and volume of the peptide is the same as the first administration, and may include administration of a solution having a lower or higher concentration and volume of the peptide. The selection of an appropriate subsequent peptide solution administration may be based on imaging the target area and the area surrounding the target area and confirming the need based on the condition of the subject. The predetermined interval may be the same or different for each subsequent administration. In some embodiments, the peptide, peptide solution, or hydrogel is prolonged at predetermined intervals to maintain at least partial pleural defect occlusion or prevention of bronchial anastomosis leakage in the subject for the life of the subject. Can be administered over a period of time. The predetermined interval may be the same or different for each subsequent administration. This may depend on whether the hydrogel formed from the previous administration is partially or completely destroyed or degraded. Subsequent administration may include administration of a solution in which the concentration and volume of the peptide is the same as the first administration, and may include administration of a solution in which the concentration and volume of the peptide are lower or higher. Selection of the appropriate subsequent administration of the peptide solution may be based on imaging or visualizing the target area and the area surrounding the target area to confirm the need based on the condition of the subject.

  Administration of the self-assembling peptide may include applying a solution containing the self-assembling peptide to the surface of the target area. In other embodiments, the solution can be applied through or within the target area. For example, rather than applying the solution to the surface of the target area, a delivery device can be positioned within the leak area to administer, eg, inject, the solution into the leak area.

  These administration procedures can be achieved by appropriately positioning the delivery device. As mentioned above, the delivery device may be a syringe. The syringe may have a specific gauge to allow for proper flow of the solution over or into the target area to achieve treatment for lung leakage.

  A further procedure for treatment may include administering a saline solution to the target area after applying the solution containing the self-assembling peptide. This increases the mechanical strength of the resulting hydrogel barrier, e.g., the storage modulus of the resulting hydrogel barrier compared to a hydrogel barrier without further treatment with saline. Due to this increase, an excellent treatment for lung leakage can be provided.

  Self-assembling peptides of the present disclosure, such as RADA16, are peptide sequences that lack distinct physiologically active or biologically active motifs or sequences and therefore may not impair endogenous cellular function. obtain. Physiologically active motifs can control many intracellular phenomena such as transcription, and the presence of physiologically active motifs allows cytoplasmic proteins or cells with enzymes that recognize the motifs. This leads to phosphorylation of surface proteins. The presence of a physiologically active motif in a peptide can activate or suppress the transcription of proteins having various functions. The self-assembling peptides of the present disclosure may lack such physiologically active motifs and therefore do not have this risk.

  Add sugar to the self-assembling peptide solution to improve the osmotic pressure of the solution from hypotonic to isotonic without reducing the treatment of pulmonary leakage, thereby increasing biological safety be able to. In a particular example, the sugar may be sucrose or glucose.

  The optimal length for film formation can vary with amino acid composition. Stabilizers contemplated by the peptides of the present disclosure are complementary peptides that maintain a constant distance between the peptide backbones. Peptides capable of maintaining a constant distance when paired are referred to herein as structurally compatible. The interpeptide distance can be calculated for each ionized or hydrogen bonded pair by taking the total number of unbranched atoms on the side chain of each amino acid in the pair. For example, lysine and glutamic acid have 5 and 4 unbranched atoms on their side chains, respectively. Peptides can be chemically synthesized or purified from natural and recombinant sources. The use of chemically synthesized peptides can allow the peptide solution to be free of unidentified components, such as unidentified components derived from the extracellular matrix of another animal or microorganism. . Therefore, due to this property, it is possible to eliminate the concern of infection including the risk of virus infection as compared with conventional tissue-derived biomaterials. This eliminates concerns of infection, including infections such as bovine spongiform encephalopathy (BSE), and makes the peptide highly safe for treating lung leakage.

  The initial concentration of the peptide can be a factor in the size and thickness of the membrane, hydrogel, or scaffold formed. Generally, the higher the peptide concentration, the higher the degree of membrane or hydrogel formation. Hydrogels, or scaffolds, formed at higher initial peptide concentrations (about 10 mg / ml) (about 1.0 w / v percent) can be thicker and therefore more potent.

  The formation of a membrane, hydrogel, composition, or scaffold is very fast, which can be on the order of seconds or minutes. The formation of the membrane or hydrogel may be irreversible. In certain embodiments, formation may be reversible. The hydrogel can be formed immediately upon administration to the target area. Hydrogel formation can occur within about 1-2 minutes after administration. In another example, formation of the hydrogel can occur within about 3-4 minutes after administration. In certain embodiments, the time taken to form the hydrogel is, at least in part, the concentration of the peptide solution, the volume of the peptide solution applied, and the conditions in the area of application or injection (eg, one time in the area of application). Concentration of valent metal cations and / or anions, pH of the area, and presence of one or more fluids at or near the area, additional components added to the solution before or after administration to the target area ) May be based on one or more of the following. The process may not be affected by a pH less than or equal to 12, and by temperature. The membrane or hydrogel can be formed at a temperature ranging from 1 to 99 degrees Celsius.

  The hydrogel can remain in the target area for a period of time sufficient to produce the desired effect using the methods and kits of the present disclosure. The desired effect of using the materials, compositions, methods and kits of the present disclosure may be to treat an area or to heal an area where a surgical procedure has been performed at or near a surgical site or where a bronchial anastomotic leak has occurred. It can be. For example, a desired effect using the materials, compositions, methods and kits of the present disclosure may be to treat an area or to assist in healing an area where lung surgery will be performed.

  The materials and methods of the present disclosure, including the use of solutions, hydrogels, compositions, or membranes comprising the self-assembling peptides described herein to treat lung leakage, provide a target for lung leakage. An area is provided for creating a hydrogel barrier. A property of a hydrogel barrier that can determine the relevance of a successful treatment is burst pressure, or burst pressure resistance. Burst pressure may refer to the pressure at which the hydrogel barrier fails. For example, this may be a pressure at which the hydrogel barrier no longer operates as desired to provide adequate treatment to the target area. The burst pressure may be the pressure at which the blockage or occlusion provided by the hydrogel barrier allows the passage of air.

In some embodiments, by using the materials and methods of the present disclosure, similar to or higher than that achieved in normal tissue (eg, undamaged tissue or tissue without leakage). It is desirable to provide a burst pressure. It is desirable to use the materials and methods of the present disclosure to provide a burst pressure similar to that exhibited by normal or average lung function. Normal, for healthy tissue, bursting pressure that is generally observed may be from about 20 to about 20 cmH ₂ O. In certain embodiments, a burst pressure of 35 cmH ₂ O or greater is a desirable or tolerable burst pressure for a hydrogel barrier to treat lung leaks (eg, pleural defects or bronchial anastomotic leaks). . In certain embodiments, the burst pressure can be increased after the step of administering a solution comprising the self-assembling peptide. For example, the burst pressure can be increased from 1 minute to 2 minutes, up to 10 minutes. In some embodiments, between about 0 minutes to one minute, it is desirable to 2 minutes 1 minute, or 2 minutes to 5 minutes burst pressure is at least 35cmH ₂ O. In certain embodiments, it is possible to achieve at least 35cmH 2 _O burst pressure during a period suitable for permitting the treatment of lung leak. This period may be appropriate for proper treatment by the clinician. Less than about 5 minutes, less than about 3 minutes, less than about 2 minutes, it is possible to achieve at least 35CmH 2 _O burst pressure of less than about less than 1 minute, or about 30 seconds.

There is an effect of the gel time after application. The burst pressure increases with time between 1 minute, 2 minutes, and 10 minutes. Two minutes may be preferred to provide a seal. Hydrogel barrier, regardless of the size of the target area, may be suitable to provide an effective burst pressure of at least 35cmH 2 _O. For example, 14g, 16g, 18g or defects were created by puncturing surface using needle 22g are not dramatically affect the burst pressure 35cmH ₂ O,.

  The period during which the membrane or hydrogel can remain in the desired area can be about 10 minutes. In certain instances, the membrane or hydrogel can remain in the desired area for about 35 minutes. In certain further examples, the membrane or hydrogel can remain in the desired area for one or more days, up to one or more weeks. In other examples, the membrane or hydrogel can remain in the desired area for up to 30 days or more. The membrane or hydrogel can remain in the desired area indefinitely. In another example, the membrane or hydrogel can remain in the desired area for a longer period of time until it is spontaneously degraded or intentionally removed. If the hydrogel degrades spontaneously over a period of time, subsequent application or injection of the hydrogel into the same or different locations can be performed.

  In certain embodiments, the self-assembling peptide can provide an enhanced effect of the self-assembling peptide, or provide another effect, treatment, therapy, or otherwise in one or more of the subjects. Can be prepared with one or more components capable of interacting with the components of. One or more other components may provide storage modulus, higher mechanical strength as measured by G ′, and improved gelation kinetics, eg, more rapid gelation into a hydrogel or hydrogel barrier. Can bring.

  For example, the pH of the self-assembling peptide in the form of a self-assembling peptide solution or composition can be adjusted. The pH of the self-assembling peptide in the form of a self-assembling peptide solution or composition can be raised or lowered. This can be performed by adjusting the pH of the self-assembling peptide solution by adding a pH adjuster. The pH adjuster can be, for example, a salt, saline solution or buffer solution. The pH adjuster can be selected based on the amino acid sequence of the self-assembling peptide, the type of salt (s), the concentration of one or more salts, and the pH of the pH adjuster. In certain embodiments, the pH of the solution comprising the self-assembling peptide is between about 2.5 and about 4.0.

  Solutions and compositions comprising the self-assembling peptides provided by the present disclosure can be prepared with additional components, for example, one or more salts. Preparation of the solution may include adding the self-assembling peptide in the form of, for example, a peptide powder or a peptide solution to the saline solution. In other embodiments, preparing the solution may include adding a salt or saline solution to the self-assembling peptide in the form of a peptide powder or peptide solution. In another embodiment, preparing a solution comprising the self-assembling peptide comprises adding water to the peptide powder of the self-assembling peptide to provide an aqueous peptide solution. The water can be deionized water or any purified water suitable for preparing a peptide solution. The water may be of a medical device acceptable grade or a pharmaceutically acceptable grade. The peptide powder and water can optionally be mixed. The salt or saline solution can then be added to the aqueous peptide solution. The salt or saline solution and the aqueous peptide solution can then be mixed.

  The saline solution can be provided for use in a solution containing the self-assembling peptide, for adding to a solution containing the self-assembling peptide, or for adding to a hydrogel or composition containing the self-assembling peptide. . The saline solution should be provided at a certain concentration, with certain anions and cations, to give the solution containing the self-assembling peptide, or the resulting hydrogel, or hydrogel barrier the desired properties. Can be. For example, the saline solution can be provided to have mechanical strength (storage modulus), stiffness, viscosity, gelation kinetics, ionic strength, pH, or burst pressure (burst pressure resistance).

  The saline solution may include monovalent and / or divalent cations and / or anions. The salt solution contains at least one cation selected from the group consisting of ammonium ion, iron ion, magnesium ion, potassium ion, pyrimidium ion, quaternary ammonium ion, sodium ion, potassium ion, and calcium ion. May include. The salt solution is a group consisting of chloride, sulfate, acetate, carbonate, chloride, citrate, cyanide, fluoride, sulfate, nitrate, nitrite, and phosphate. It may include at least one anion selected from the group consisting of:

  In some embodiments, the saline solution comprises at least one of calcium chloride, sodium chloride, and potassium chloride.

In certain embodiments, the solution comprising the self-assembling peptide may comprise (RADA) ₄ (SEQ ID NO: 1) at a concentration of about 0.5 w / v percent. The solution may further include a calcium chloride concentration of about 0.125M. The solution may further provide a storage modulus of about 25 Pa.

In certain embodiments, the solution comprising the self-assembling peptide may comprise (RADA) ₄ (SEQ ID NO: 1) at a concentration of about 0.5 w / v percent. The solution may further include a calcium chloride concentration of about 0.250M. The solution may further provide a storage modulus of about 44 Pa.

In certain embodiments, the solution comprising the self-assembling peptide may comprise (RADA) ₄ (SEQ ID NO: 1) at a concentration of about 0.5 w / v percent. The solution may further include a calcium chloride concentration of about 0.500M. The solution may further provide a storage modulus of about 52 Pa.

In certain embodiments, the solution comprising the self-assembling peptide may comprise (RADA) ₄ (SEQ ID NO: 1) at a concentration of about 2.5 w / v percent. The solution may further include a calcium chloride concentration of about 0.125M. The solution may further provide a storage modulus of about 600 Pa.

  In embodiments, the solution comprising the self-assembling peptide may have a salt concentration between about 0.005M and about 1M. In certain embodiments, the solution comprising the self-assembling peptide may have a salt concentration between about 0.125M and about 0.500M. In certain embodiments, the solution comprising the self-assembling peptide may have a salt concentration of about 0.25M.

In embodiments, the solution comprising the self-assembling peptide may comprise an isotonic solution or may be supplemented with an isotonic solution. The isotonic solution may be for a subject, for example, a body fluid of the subject or a local physiological condition in a target area. The isotonic solution may include at least one of sodium chloride, potassium chloride, calcium chloride and water. This solution may contain hydrochloric acid or sodium hydroxide, which can be used to adjust the pH. To prepare this solution, 8.6 g of NaCl, 0.3 g of KCl, 0.33 g of CaCl ₂ can be dissolved in 1 liter of distilled water. The pH of this solution may be about 5.4. The pH of this solution can be adjusted using an acid or a base, or a pH adjuster. The pH adjuster may be sodium bicarbonate. This solution can be referred to as Ringer's solution.

  In embodiments, the solution containing the self-assembling peptide may include a contrast agent or may have a contrast agent added. The contrast agent can be used to visualize a solution or hydrogel or hydrogel barrier containing the self-assembling peptide. The contrast agent can provide or convince the physician of the location of the solution or hydrogel or hydrogel barrier containing the self-assembling peptide. The contrast agent may include at least one of a sulfate ion and a sodium ion.

In embodiments, various self-organizations include, but are not limited to, (RADA) ₄ (SEQ ID NO: 1), (IEIK) ₃ I (SEQ ID NO: 2), (KLDL) ₃ (SEQ ID NO: 3) The properties of the peptides can be enhanced by maintaining their salt concentration below the critical ionic strength level before the salts begin to precipitate. The critical ionic strength level of the salt will vary depending on the endogenous amino acid properties and composition of each peptide. Peptides can be dissolved in water with various salts instead of pure water to maintain the ionic strength of their salts below the critical ionic strength level before the salts begin to precipitate.

  This beneficially imparts relatively stiff properties or higher mechanical strength to the self-assembled peptide solutions and hydrogels at various salt concentrations, compared to peptide hydrogels which maintain them at zero salt concentration levels. It can be suitable for a wide range of applications. Similarly, this may result in environmental conditions when the peptide solution is administered to a target area of a subject, e.g., ionic strength at which environmental salt ionic strength exceeds the critical ionic strength before salt precipitation, such as physiological ionic strength. , The rapid gelation kinetics from the peptide solution to the peptide hydrogel can also be beneficially imparted.

According to one or more aspects, various (including but not limited to) (RADA) ₄ (SEQ ID NO: 1), (IEIK) ₃ I (SEQ ID NO: 2), (KLDL) ₃ (SEQ ID NO: 3) The properties of peptide hydrogels are enhanced by maintaining their pH levels at a high value of about 3.5 or less, while maintaining their salt concentration below the critical ionic strength level before salt precipitation. be able to.

In some embodiments, the pH of the solution comprising (RADA) ₄ (SEQ ID NO: 1) is about 3.5. In some embodiments, the solution comprising (KLDL) ₃ (SEQ ID NO: 3) has a pH of about 3.5. In some embodiments, the pH of the solution comprising (IEIK) ₃ I (SEQ ID NO: 2) is about 3.7.

  In some embodiments, a buffer, such as a buffer solution, can be added to the self-assembling peptide solution or the self-assembling peptide.

  The buffer may be an aqueous solution consisting of a mixture of a weak acid and its conjugate base, or vice versa. The pH of the buffer hardly changes with the addition of small or moderate amounts of strong acids or bases, and thus the buffer can be used to prevent changes in the pH of the solution. Buffer solutions are used as a means of maintaining the pH at a nearly constant value in a wide variety of chemical applications and are applicable to the self-assembling peptides and self-assembling peptide solutions and compositions disclosed herein. is there.

  The buffer may include at least two salts. The buffer may have a pH of about 7.4, such as, for example, a PBS buffer (phosphate buffered saline). The buffer may have a pH of about 7.2, such as a DMEM buffer. In some embodiments, the buffer may be an alkaline buffer.

In some embodiments, the solution or composition of the self-assembling peptide may be buffered with at least one of about 0.15 M sodium chloride, potassium chloride, and calcium chloride. When the self-assembling peptide is (RADA) ₄ (SEQ ID NO: 1), the buffer may contain between about 0.6M to about 1.2M salt. Where the self-assembling peptide is (IEIK) ₃ I (SEQ ID NO: 2), the buffer may comprise between about 0.6M to about 1.2M salt. When the self-assembling peptide is (RADA) ₄ (SEQ ID NO: 1), the buffer may include between about 0.02M to about 0.04M salt. Where the self-assembling peptide is (KLDL) ₃ (SEQ ID NO: 3), the buffer may include between about 0.1 M to about 0.4 M salt.

  In certain embodiments, a therapeutic method is provided that further comprises selecting a salt to provide a predetermined mechanical strength to the solution. The method may further include selecting a salt concentration to provide a predetermined mechanical strength to the solution. The method may include selecting a salt to provide a predetermined ionic strength to the solution. The method may further include the step of selecting a salt concentration to provide a predetermined ionic strength to the solution. The method may include selecting a salt to bring the solution to a predetermined pH. The method may further include selecting a salt concentration to bring the solution to a predetermined pH.

  An additional peptide comprising one or more biologically active or physiologically active amino acid sequences or motifs can be included as one of the components together with the self-assembling peptide. Other components can include biologically active compounds that can provide some benefit to a subject, such as a drug or other treatment. For example, a cancer therapeutic or anticancer drug can be administered with a self-assembling peptide, or can be administered separately.

  The peptide, peptide solution, or hydrogel may include a small molecule drug to treat the subject or prevent hemolysis, inflammation, and infection. Small molecule drugs include glucose, saccharose, purified saccharose, lactose, maltose, trehalose, dextran, iodine, lysozyme chloride, dimethylisopropylazulene, tretinoin, tocopheryl, tocopheryl, popidone iodine, alprostadil alfadex Isoamyl salicylate, α, α-dimethylphenylethyl alcohol, bacdanol, helional, silver sulfadiazine, sulphadiazine silver, bucladecine sodium, alprostadil alphadex, gentamicin sulfate, tetracycline hydrochloride, sodium fusidinate, Mupirocin calcium hydrate and It may be selected from the group consisting of Ikikosan isoamyl. Other small molecule drugs may be contemplated. Protein-based drugs may be included as administered components, including erythropoietin, tissue-type plasminogen activator, synthetic hemoglobin and insulin.

  Components can be included to protect the self-assembling peptide, solution containing the self-assembling peptide, or composition from rapid or immediate formation into a hydrogel. This can include an encapsulated delivery system that can be degraded over time to allow time-controlled release of the peptide solution to the target area and allow the hydrogel to form over a desired predetermined period of time. Biodegradable biocompatible polymers such as ethylene vinyl acetate, polyanhydrides, polyglycolic acid, collagen, polyorthoesters, and polylactic acid can be used.

  Any of the components described herein can also be included in a self-assembling peptide, a solution, composition, or kit containing the self-assembling peptide, a self-assembling peptide, a solution, containing the self-assembling peptide, a composition. It can also be administered separately from the product or kit. Furthermore, any of the methods provided and facilitated herein may be performed by one or more parties.

The peptides, peptide solutions, compositions or hydrogels of the present disclosure can be provided in a kit for treating lung leakage (eg, pleural deficiency or bronchial anastomosis). The kit may be for treating lung leakage in a subject. Instructions for administering the solution self-assembling peptide to the target area of the lung leak in the subject can also be provided in the kit. The self-assembling peptide may include an effective amount of about 7 amino acids to about 32 amino acids to form a hydrogel barrier to allow for treatment of lung leakage. In some embodiments, the self-assembling peptide may comprise, consist of, or consist essentially of about 12 amino acids to about 16 amino acids. The self-assembled peptide may comprise (RADA) ₄ (SEQ ID NO: 1), (IEIK) ₃ I (SEQ ID NO: 2), (KLDL) ₃ (SEQ ID NO: 3), and may consist essentially of: It may consist of it. The concentration of the self-assembling peptide in the solution can be any of the concentrations disclosed herein.

  Instructions for administering the solution include administering the peptide, peptide solution, or hydrogel provided herein at a dose, volume or concentration, or dosing schedule, for example, by the routes of administration described herein. Method may be included. The peptide may be amphiphilic, and at least a portion of the peptide may be one in which hydrophobic amino acids and hydrophilic amino acids alternate.

  The kit can provide the self-assembling peptide as one of a solution containing the self-assembling peptide and a powder prepared as a solution containing the self-assembling peptide. Instructions for preparing a solution containing a self-assembling peptide having an effective concentration to form a hydrogel barrier under physiological conditions to allow treatment of lung leakage can also be provided.

  The kit may also include informational materials. The informational material may be instructional, educational, marketing, or other material related to the methods described herein. In one embodiment, the informational material is the preparation of a peptide, peptide solution, or hydrogel as disclosed herein, the physical properties, concentration, volume, size, size, dimensions, of the peptide, composition, peptide solution or hydrogel. It may include information about the expiration date, and the batch or production location.

  The kit may also optionally include a device or material to allow administration of the peptide or peptide solution to a desired area. For example, a syringe, pipette, catheter, or other needle-based device can be included in the kit. Additionally or alternatively, the kit may include a guidewire, endoscope, or other ancillary equipment to effect selective administration of the peptide solution to the target area.

  The kit may additionally or alternatively include other components or components, such as peptide solutions, hydrogels or components that may assist in placing the scaffold. The kit can provide instructions for combining a sufficient amount or volume of the peptide solution with a sucrose solution that may or may not be provided with the kit. Instructions for diluting the peptide solution to administer an effective concentration of the solution to the target area can be provided. The instructions may describe diluting the peptide solution with a diluent or solvent. The diluent or solvent may be water. Instructions for determining at least one of an effective concentration of the solution and an effective amount of the solution for the target area can be further provided. This may be based on various parameters discussed herein and may include bronchial anastomosis leakage or wound lesion or site diameter at the target area.

  Other components or components can be included in the kit in the same or a different composition or container as the peptide, peptide solution, or hydrogel. As one or more components, one or more components of the subject can provide an enhanced effect of the self-assembling peptide, or provide another effect, treatment, therapy or otherwise. And components capable of interacting with. For example, an additional peptide containing one or more biologically active sequences or physiologically active sequences or motifs can be included as one of the components together with the self-assembling peptide. Other components may include biologically active compounds that may provide some benefit to the subject, such as a drug or other treatment. For example, a cancer therapeutic or anticancer drug can be administered together with the self-assembling peptide, or can be administered separately. The peptide, peptide solution, or hydrogel may include a small molecule drug for treating a subject or preventing hemolysis, inflammation, and infection, as disclosed herein. A sugar solution such as a sucrose solution can be provided with the kit. The sucrose solution may be a 20% sucrose solution.

  Other components disclosed herein throughout this disclosure can also be included in the kit. For example, the kit may further include a saline solution separately from or in combination with the self-assembling peptide. The kit may further comprise, for example, a sugar or a sugar solution, such as sucrose, provided separately from or together with the self-assembling peptide. Instructions for combining one of the solution or peptide powder containing the saline solution and the self-assembling peptide can be provided. The kit may further comprise an isotonic solution or a contrast agent as added to the self-assembling peptide solution or powder or as part of the self-assembling peptide solution.

  In some embodiments, the components of the kit are stored in a sealed vial having, for example, a rubber or silicone lid (eg, a polybutadiene or polyisoprene lid). In some embodiments, the components of the kit are stored under inert conditions (eg, under another inert gas such as nitrogen or argon). In some embodiments, the components of the kit are stored under anhydrous conditions (eg, using a desiccant). In some embodiments, the components of the kit are stored in a light-tight container, such as an amber vial.

  As part of or separate from the kit, a syringe or pipette can be pre-filled with a peptide, peptide solution, or hydrogel disclosed herein. Provide the user with a syringe or pipette with the self-assembling peptide solution with or without the use of other devices, and the syringe or pipette with the self-assembling peptide solution with or without the use of other devices. Provides a method for directing administration to a target area. Other devices may include, for example, a catheter with or without a guidewire.

  The self-assembling peptide of the kit can be any of the peptides provided in the present disclosure, and any of the components described in the present disclosure, such as various salts, pH adjusters, buffers, alkaline buffers Can be provided in the kit, together with the self-assembling peptide in the kit, or separately from the self-assembling peptide in the kit.

In embodiments, a self-assembling peptide comprising an effective amount and effective concentration of about 7 to 32 amino acids for use in forming a hydrogel barrier under physiological conditions to treat lung leakage. There is provided a composition comprising: The hydrogel barrier composition can result in rupture pressure resistant of at least 35H 2 _O. The self-assembling peptide of the composition can be selected from the group consisting of (RADA) ₄ (SEQ ID NO: 1), (IEIK) ₃ I (SEQ ID NO: 2), and (KLDL) ₃ (SEQ ID NO: 3). Effective concentrations to enable treatment of lung leakage include self-assembling peptide concentrations ranging from about 0.1 weight / volume (w / v) percent to about 3 w / v percent. The composition may be substantially free of cells. The composition may be substantially free of drug. The composition may further include any one or more of the components disclosed herein. For example, the composition can include any one or more of the cations, anions, salts, buffers, contrast agents, isotonic solutions, pH adjusters, and sugars disclosed herein. May be included at various concentrations as disclosed in the publication. The composition may have properties such as mechanical strength, pH, gelation kinetics, and ionic strength disclosed herein. The composition can be used to treat lung leakage (eg, pleural deficiency or bronchial anastomosis leakage) and can be a treatment for a subject such as a mammal or human.

  Methods can also be provided that facilitate treatment of lung leakage in a subject. The method comprises providing a solution comprising an effective amount and an effective concentration of a self-assembling peptide comprising from about 7 amino acids to about 32 amino acids to enable prevention of lung leakage and providing a hydrogel barrier under physiological conditions. And providing instructions to administer the solution to the target area of the pulmonary system by introducing the solution through a delivery device located within the leak of the lung.

  The method may further comprise the step of providing instructions for visualizing an area comprising at least a portion of a lung leak disclosed herein. Instructions for visualizing an area that includes at least a portion of a lung leak, the method comprising: identifying a target area of the pulmonary system; introducing a delivery device; placing an end of the delivery device within the target area; Instructions can also be provided that include at least one of administering a solution; removing the delivery device from lung leakage; and monitoring lung leakage after removing the delivery device. Instructions for visualizing the area may be provided within about 1 minute to about 5 minutes after the step of administering the solution. The method may further include providing instructions for preparing an effective amount and / or concentration based, in part, on the dimensions of the target area of lung leakage, as discussed in the present disclosure.

The self-assembling peptide is selected from the group consisting of (RADA) ₄ (SEQ ID NO: 1), (IEIK) ₃ I (SEQ ID NO: 2), and (KLDL) ₃ (SEQ ID NO: 3). The method may further include providing instructions for monitoring an area around the target area. The method may further include providing a solution and instructions for use after the surgical procedure. Disclosed herein is a method comprising providing a solution comprising a self-assembling peptide having an effective concentration to form a hydrogel barrier under physiological conditions to allow for prevention of lung leakage. Providing instructions for preparing the peptide solution being prepared.

(Example 1)
Effect of pH Level on Rheological Properties of Peptide Hydrogels The effect of Dulbecco's Modified Eagle's Medium (DMEM) (pH 7.4) on the rheological properties of IEK13, KLD12, and PuraMatrix® was measured using a rheometer with a 40 mm plate (see Table 1). AR500, TA Instruments). DMEM is generally a cell culture containing 6.4 g / L NaCl, 3.4 g / L NaHCO ₃ (sodium bicarbonate), trace amounts of other salts, various amino acids, and 4.5 g / L glucose. It is a medium. The pH level of DMEM is generally 7.2 ± 0.2 and the osmolality is 335 ± 30 mOsm / KgH ₂ O; both measurements are close to human physiological fluids such as blood.

  Testing was performed after maintaining the peptide solution (1%) at 4 ° C. for at least 48 hours. To perform the experiment, 1 mL of the peptide solution was gently pipetted and placed on the plate of the rheometer. 2 mL of DMEM solution was gently added around the peptide solution. The peptide solution was treated with DMEM for 2 minutes, then the medium was removed and the plate was placed in a measurement geometry gap of approximately 450 μm. The measurement was performed at 37 ° C. after a relaxation time of 2 minutes. The frequency test was performed from 1 rad / s to 100 rad / s at a vibration stress of 1 Pa.

  As shown in FIG. 1A, the rheological properties of the peptides (1%) were compared before and after DMEM treatment for 2 minutes. The fold increase in storage modulus after DMEM treatment for 2 minutes is shown in FIG. 1B. Each of the peptides showed a large increase in storage modulus after DMEM treatment. The fold difference between the storage modulus after DMEM treatment between PuraMatrix®, KLD12 and IEIK13 was relatively small compared to that before DMEM treatment. Similarly, after DMEM treatment, the fold increase in storage modulus exhibited by the more rigid peptide solution (ie, IEIK13) was lower than that of the weaker peptide solution (ie, PuraMatrix®). This observation suggests that after DMEM treatment, critical intermolecular interactions occur that determine the final stiffness after DMEM treatment.

(Example 2)
Optimization of the pH Level of the Peptide Solution As an example, to adjust the pH level of the peptide solution, 0.1 N NaOH was added to 2 mL of the 2.5% peptide solution and their pH and appearance were measured. The results are shown in Table 1. In particular, increasing the pH to approximately 3.5 or less did not change the clear color of the PuraMatrix®, IEIK13, and KLD12 solutions, but increased their apparent stiffness.

(Example 3)
Rheological Properties of pH-Adjusted Peptide Solutions Based on visual observation of the effect of pH levels on the properties of peptide solutions, the pH levels of peptide solutions were adjusted to 3.4 (PuraMatrix® and KLD12) or 3.7 ( After adjusting to IEIK13), the effect on the rheological properties of the peptide solution was evaluated. If the pH level of the peptide solution is higher than 3.5 (PuraMatrix® and KLD12) or 3.7 (IEIK13), the peptide solution will begin to phase separate and become cloudy. The rheological properties of the PuraMatrix® solution, KLD12 solution and IEIK13 solution were higher at pH 3.4. The results are shown in FIG. 2 for KLD12 1%, FIG. 3 for IEIK13 1%, and FIGS. 4-5 for PuraMatrix® 1% and 2.5%, respectively. The stress sweep test was performed at 10 rad / s. The frequency sweep test was performed at 1 Pa.

(Example 4)
Additional Rheological Properties of the pH-Adjusted Peptide Solution The effect on the rheological properties of the peptide solution after adjusting the pH level of the peptide solution to 3.4 (PuraMatrix® and KLD12) or 3.7 (IEIK13). Based on the results, the effect on the rheological properties of the peptide solution at various pH levels was evaluated. The rheological properties of the PuraMatrix® and IEIK13 solutions increase as the pH is adjusted to 3.4. The rheological properties of the peptides were evaluated at various concentrations using a rheometer (DHR-1, TA Instruments) with a 20 mm plate. The results are shown in FIG. 6A for a PuraMatrix® 2.5% solution and in FIG. 6B for a 1.5% IEIK13 solution, respectively. A frequency sweep test was performed at 1 Pa from 1 rad / sec to 10 rad / sec, and the storage modulus at 1 rad / sec was selected for the data.

(Example 5)
Effect of pH level on rheological properties of peptide hydrogel at various concentrations before / after DMEM treatment Based on the results of the effect on the rheological properties of peptide solution after adjusting the pH level of peptide solution, The effect of the different pHs on the rheological properties of the peptide hydrogels afterwards was evaluated and compared with the effect on the rheological properties of the peptide solutions at different pHs before DMEM treatment. The rheological properties of PuraMatrix® hydrogels and IEIK13 hydrogels after DMEM treatment increase with pH adjustment up to 3.4. The results are shown in FIGS. 7A-7B for PuraMatrix® and IEIK (SEQ ID NO: 5) in FIGS. 8A-8B, respectively. A frequency sweep test was performed at 1 Pa from 1 rad / sec to 10 rad / sec, and the storage modulus at 1 rad / sec was selected for the data.

(Example 6)
Effect of pH-Adjusted Peptide Hydrogels on Gelation Kinetics To identify pH levels optimized for the peptides described herein, the effect of pH levels on the nature of gelation kinetics was evaluated. The rapid gelation kinetics of PuraMatrix® and other peptides in bodily fluids can generally improve their function and response time for various clinical applications. The pH level can provide a response time before gelation is initiated when treated with simulated body fluids, including but not limited to DMEM. The unadjusted (pH 2.2) PuraMatrix® did not show an increase in storage modulus over the first 13 seconds, whereas the pH adjusted PuraMatrix® caused rapid gelation. The resulting immediate increase in storage modulus was shown. The rapid response time of PuraMatrix® and other peptides in bodily fluids can generally improve their function and response time for various clinical applications.

  The time sweep test was performed at 1 rad / sec and 1 Pa using a 20 mm plate and a gap distance of 500 μm. During the time sweep test of the PuraMatrix® 2.5% solution, DMEM was added to the chamber around the measurement plate at time 0 to immerse the PuraMatrix® solution. The results are shown in FIG. 9A for a PuraMatrix® 2.5% solution.

  The unmodified pH IEK (SEQ ID NO: 5) showed an immediate increase in storage modulus, whereas the unregulated (pH 2.2) PuraMatrix® stored for the first 13 seconds. No increase in modulus was shown. The pH adjusted IEIK13 also showed an immediate increase in storage modulus due to rapid gelation. The rapid response time of IEK13 in bodily fluids can generally improve its function and response time for various clinical applications.

  The time sweep test was performed at 1 rad / sec and 1 Pa using a 20 mm plate and a gap distance of 500 μm. During the time sweep test of the IEIK13 1.5% solution, DMEM was added into the chamber around the measurement plate at time 0, soaking the IEIK13 1.5% solution and recording data continuously. The results are shown in FIG. 9B for the IEIK13 1.5% solution.

(Example 7)
Effect of Salt Ionic Strength Level on Peptide Solutions and Hydrogels To identify salt ionic strength levels optimized for the peptides described herein, the effect of salt ionic strength levels on the properties of the peptide solution was evaluated. Increasing the salt ionic strength levels of PuraMatrix® and other peptides can generally improve their function and mechanical strength for various clinical applications. By way of example, to adjust the salt ionic strength of the peptide solution, various salt buffer solutions including NaCl, KCl, MgCl ₂ , CaCl ₂ and DPBS (10 ×) were added to 2 mL of the 1.5% peptide solution.

The results for PuraMatrix® are shown in Table 2a. In particular, there was no significant change in the clear color of the PuraMatrix® solutions by increasing the salt ionic strength by approximately 0.85 to 1.15M (depending on the various salts), but their apparent stiffness was Increased. The results for KLD12 are shown in Table 2b. In particular, increasing the salt ionic strength to approximately 0.25-0.35M (depending on the various salts) did not significantly change the clear color of the KLD12 solutions, but increased their apparent stiffness. The results for IEIK13 are shown in Table 2c. In particular, increasing the salt ionic strength (approximately 0.025-0.035M) (depending on the various salts) did not change the clear color of the IEK13 solutions, but did increase their apparent stiffness.

  The results from Tables 1a-1c show that the critical salt ionic strength at which the three peptides begin to become cloudy is shown as follows: PuraMatrix® (0.9-1.2M)> KLD13 (0.3 ０．４0.4M)> IEIK13 (0.03 to 0.04M).

(Example 8)
Effect of salt ionic strength level on the rheological properties of peptide solution Based on visual observation of the effect of salt ionic strength on the properties of peptide solution, the ionic strength level of peptide solution can be adjusted using NaCl to make each peptide cloudy The effect on the rheological properties of the peptide solution after adjustment to 0.7M (PuraMatrix®), 0.2M (KLD12) or 0.02M (IEIK13) slightly below the critical ionic strength was evaluated. If the ionic strength level of the peptide solution using NaCl is higher than 0.9 M (PuraMatrix®), 0.3 M (KLD12) or 0.03 M (IEIK13), the peptide solution begins to phase separate and becomes cloudy. , Become weak. The rheological properties of the PuraMatrix®, KLD12 and IEIK13 solutions are based on the ionic strength level of the peptide solution at 0.7 M (PuraMatrix®), 0.2 M (KLD12) or 0.2 M with NaCl. It was higher after adjusting to 02M (IEIK13). The results are shown in FIG. 10 for KLD12 1%, FIG. 11 for IEIK13 1% and FIG. 12 for PuraMatrix® 1%, respectively. A frequency sweep test was performed at 1 Pa from 1 rad / s to 10 rad / s.

(Example 9)
Further Effect of Salt Ionic Strength Level on Rheological Properties of Peptide Solution Based on the results of the effect on the rheological properties of the peptide solution, the ionic strength level of the peptide solution was increased to 0.7 M with NaCl (PuraMatrix®). , 0.2M (KLD12) or 0.02M (IEIK13), the effect on rheological properties of the peptide solution at various salt ionic strengths was evaluated. The rheological properties of the PuraMatrix® 1% solution increase with adjustment of the ionic strength up to 0.7M, but decrease above 0.7M. The rheological properties of the IEIK13 1% solution increase with adjustment of the ionic strength up to 0.03M, but decrease with adjustment above 0.03M. These results are in good agreement with visual inspection of peptide solutions of various salt ionic strengths. The results are shown in FIG. 13 for the PuraMatrix® 1% solution and in FIG. 14 for the IEIK13 1% solution. A frequency sweep test was performed at 1 Pa from 1 rad / sec to 10 rad / sec, and the storage modulus at 1 rad / sec was selected for the data.

(Example 10)
Effect on the Rheological Properties of the Peptide Solution after DMEM Treatment Based on the results of the effect on the rheological properties of the peptide solution after adjusting the ionic strength level of the peptide solution, the rheological properties of the peptide hydrogel after 10 minutes of DMEM treatment The effect on properties was evaluated. The rheological properties of the PuraMatrix (R) hydrogel after DMEM treatment increased up to 0.7 M in ionic strength adjustment, but declined above 0.7 M. The rheological properties of the IEIK13 hydrogel after DMEM treatment did not change significantly until the ionic strength adjustment was 0.025M, but decreased when the ionic strength adjustment exceeded 0.03M. When the PuraMatrix® solution exceeds 0.9 M NaCl ionic strength at which it begins to become cloudy, the rheological properties of PuraMatrix® do not change with the DMEM treatment, thereby demonstrating that no gelation has occurred. You. The results are shown in FIG. 15 for PuraMatrix® 1% hydrogel and in FIG. 16 for IEIK13 1% hydrogel, both after DMEM treatment for 10 minutes. A frequency sweep test was performed at 1 Pa from 1 rad / sec to 10 rad / sec, and the storage modulus at 1 rad / sec was selected for the data.

(Example 11)
Effect of Various Salts Based on the effect on the rheological properties of peptide solutions and hydrogels after adjusting the ionic strength levels of peptide solutions and hydrogels with NaCl, various salts (KCl, MgCl ₂ , and The effect of CaCl ₂ ) was also evaluated. The rheological properties of the PuraMatrix® solution increase for all salts when the ionic strength is adjusted to 0.15M. The enhancement of the rheological properties of the PuraMatrix® solution with different salts did not differ significantly. However, the increase in rheological properties of the PuraMatrix® solution can vary depending on the salting out constant, K, of each salt. The constant K is a constant in the Cohen equation: logS = B-KI, where S is the solubility, B is the ideal solubility, K is the salting-out constant, and I is the ionic strength. is there. As the value of the constant K and the ionic strength of the salt increase, the solubility of the peptide decreases, resulting in strong peptide self-assembly with increased hydrophobicity effects and higher rheological properties of the peptide solution. . The constant K of NaCl may be higher than other salts. Thus, the rheological properties of the PuraMatrix® solution with NaCl were slightly higher than those with KCl and CaCl ₂ . The rheological properties of the PuraMatrix® hydrogels after 10 minutes of DMEM treatment were also evaluated using various salts and adjusting the ionic strength, and the results were evaluated using various salts ((NaCl, KCL, MgCl ₂ , and CaCl ₂ ), ionic strength of 0.15 M) was comparable to the increase in rheological properties of the PuraMatrix® solution. The results are shown in FIG. 17 for the PuraMatrix® 1% solution before DMEM treatment and in FIG. 18 for the PuraMatrix® 1% hydrogel after DMEM treatment for 10 minutes. A frequency sweep test was performed at 1 Pa from 1 rad / sec to 10 rad / sec, and the storage modulus at 1 rad / sec was selected for the data. * Indicates that the data is significantly higher than the PuraMatrix® control data (P <0.05). # Indicates that the data is significantly lower than the PuraMatrix® 1% NaCL 0.15M (ionic strength) data (P <0.05).

(Example 12)
Influence of various salts on gelation kinetics To identify the possibility of peptide gelation when the peptide solution was placed in an environment with a high salt ion strength level, the salt ion strength level around the peptide solution was compared with the properties of the peptide solution. The impact was evaluated. For example, the hydrogel can be placed in an isotonic body fluid corresponding to a saline buffer (0.15 M NaCl). As demonstrated previously, self-assembling peptides, including but not limited to PuraMatrix®, KLD12 and IEIK13, form hydrogels when treated at neutral pH. The effect of saline treatment on the gelation of the peptide solution without the effect of pH was evaluated. When the peptide solutions were treated with saline buffer, their pH did not change. After treatment with saline buffer, only IEIK13 showed rapid gelation, whereas PuraMatrix® and KLD13 showed no or negligible gelation. This is due to the fact that IEK13 is much more sensitive to salt ionic strength levels. The rapid gelation of IEIK13 at a salt ionic strength level similar to that of body fluids and isotonic salts can generally improve its function and gelation rate for various clinical applications. The results for the IEIK13, KLD12 and PuraMatrix® solutions are shown in FIG. The time sweep test was performed at 1 rad / sec and 1 Pa using a 20 mm plate and a gap distance of 500 μm. During the time sweep test of the IEIK13 1.5% solution, the KLD12 1.5% solution, and the PuraMatrix® 2.5% solution, DMEM was added to the chamber around the measurement plate at time 0 to add PuraMatrix ( (Registered trademark) solution.

(Example 13)
Effect of Adjusting Salt Ionic Strength and pH on Rheological Properties According to one or more embodiments, IEIK13, KLD12, and PuraMatrix® can be used in salt buffers such as NaCl, and salt ionic strength in critical salt. Can be dissolved at elevated pH levels adjusted using an alkaline salt buffer such as NaOH to maintain the pH below about 2.5 to 4.0. It may have more rigid properties. For PuraMatrix®, KLD13 and IEIK13, the peptide solution is still clear using 0.9% NaCl (ionic strength: 0.15 M) at pH 3.4 adjusted with NaOH. The rheological properties of PuraMatrix® using 0.9% NaCl (ionic strength: 0.15M) at pH 3.4 were stronger than those of PuraMatrix control and PuraMatrix using only 0.9% NaCl. The effect of adjusting the salt ionic strength and pH on the rheological properties of a PuraMatrix® 2.5% solution is shown in FIG. A frequency sweep test was performed at 1 Pa from 1 rad / sec to 10 rad / sec, and the storage modulus at 1 rad / sec was selected for the data.

(Example 14)
Effect of cations In the rheological comparison of 2.5% RADA16 and 2.5% RADA16 + NaCl, KCl, and CaCl2, 0.005M, 0.05M, 0.125M, 0.25M, 0.5M, and 1M A solution of 0.5% RADA16 mixed with NaCl, KCl, and CaCl2 was prepared. To observe the effects of cations, sodium ions (Na +), potassium ions (K +), and calcium ions (Ca2 +), anions, chloride ions (Cl-) were also maintained. FIG. 21 provides a basic understanding of how cation fluctuations in saline solutions affect the viscoelasticity and stiffness of self-assembling peptides. Ca provided the best stiffness enhancement compared to either the same molarity of Na or K. This should be due to the ionic strength of Ca being four times that of Na and K at the same molarity. Thus, as shown in FIGS. 17-18 and Tables 2a-c, the effect of salts on peptide solutions is more related to their ionic strength but not their molarity. In some embodiments, there is a correlation between the nature of the peptide solution and the salt based on the concentration of the salt.

(Example 15)
Mechanical Strength Rheological measurements of stiffness of 2.5% RADA16 and 2.5% RADA16 + 0.25M CaCl2 were evaluated. FIG. 22 compares the stiffness of a highly concentrated solution of RADA16 with another highly concentrated solution of RADA16 supplemented with 0.125 M CaCl2 and provides a basic understanding of the viscoelasticity of peptides and peptide mixtures. The addition of the cationic solution resulted in a noticeable increase in the stiffness between the two solutions. Ca was shown to provide mechanical enhancement of RADA16, even at high concentrations, using the optimal concentration range.

(Example 16)
Reversibility Rheological measurements of reversibility of a 0.5% RADA16 solution with +0.125 M, 0.25 M, and 0.5 M CaCl 2 were evaluated. Solutions of 0.5% RADA16 mixed with 0.125M, 0.25M, and 0.5M CaCl2 were prepared. The structure of the self-assembled peptide solution was thoroughly disrupted by applying mechanical stress by vortexing and sonication. The mixture was left at room temperature for 48 hours to allow self-assembly. FIGS. 23A-23B present the basic viscoelasticity of the peptide mixture, and the salt was removed as demonstrated by the significant difference between the 2.5% RADA16 + 0.5 M CaCl 2 control and the perturbed sample. It shows that the reversibility of the associated peptide solution can be controlled and maintained even after disturbing the structure. Mixtures within the optimal salt concentration range remained reversible. * Indicates significantly different control and perturbed samples. FIG. 23a shows the raw rheological data of a control peptide solution with salt and a peptide solution with a disturbed salt, and FIG. 23b presents a comparison of the rigidity of the control peptide solution with the disturbed peptide solution. It is.

(Example 17)
Gelation Kinetics Rheological measurements of the gelation kinetics of 0.5% RADA16 + NaCl, KCl, and CaCl2 were evaluated. Gelation kinetics were observed by preparing a solution of 0.5% RADA16 and treating with some cations (eg, Na, Cl, K) and anions (eg, Cl, CO3, PO4, SO4). . The time taken to gel the peptide mixture and how to control the gel time by varying the cation / anion type and concentration were determined. Chlorine showed the fastest gelation, and sulfate showed the slowest gelation. Qualitative experiments and results obtained in vivo and in vitro supported these conclusions.

(Example 18)
Various cations Peptide hydrogel mixed with cation / anion solution that affected mechanical properties, and another peptide mixed with very low concentration of contrast agent that did not affect mechanical properties Both hydrogels were designed. The two gels comprise (1) a combination of a self-assembling peptide and Ringer's solution (pH 5.3), a well-known cation / anion solution used in the medical field, and (2) a self-assembling peptide, It was a combination of a well-known contrast agent, indigo carmine, a dye solution containing sulfate ions (anions) and sodium ions (cations). Indigo carmine contains sodium indigo disulfonate sodium (C ₁₆ H ₈ N ₂ Na ₂ O ₈ S ₂ ), water, and sodium citrate (C ₆ H ₈ O ₇ ) to adjust the pH. I do. Indigo carmine powder was used to prepare a 1% solution for use in experiments. This corresponds to 10 mg per ml of water. The concentration of the indigo carmine solution used in the experiment was 0.00585% in water.

  A 1% solution in deionized (DI) water was prepared using indigo carmine powder. Using IEIK13 powder, a 2 percent solution was prepared using DI water. The amount of IEIK was weighed and an appropriate amount of DI water was added by gently flowing to the side of the container. Mixing was achieved by vortexing for about 30 seconds and then sonicating for 30 seconds. The solution was then centrifuged at 3000 ppm for about 10 to about 15 minutes. The solution can be further vortexed and centrifuged until the solution is clear and free of bubbles.

  To obtain a final concentration of 0.00585% indigo carmine, add the required amount of 1% IC to the appropriate volume of DI water and dilute the 2% IEIK to 1.5% IEIK. The solution is then vortexed for about 30 seconds and centrifuged at 3000 rpm for about 10 to about 15 minutes. The solution can be further vortexed and centrifuged until the solution is clear and free of bubbles.

  The solution was left at room temperature overnight before use. The cap can be removed from the container to allow for more efficient foam removal during preparation.

  A rheological comparison of these mixtures and a visualization of the gel can be observed in FIGS. A stiffer gel was obtained with faster gelation kinetics where reversibility was maintained. Another gel was obtained that retained stiffness, reversibility, and gelation kinetics, but allowed tissue death due to histology. The concentrations of the contrast agent or cation / anion mixture and the mixed peptide hydrogel were based on the understanding of the use of cations and anions described herein. From the data on these RADA16 + Ringer's and IEIK13 + indigo carmine adapted peptide hydrogels, these controlled self-assembled hydrogels were rendered isotonic, injectable gels and not mechanically enhanced for visualization. It is shown that the gel can be adapted to meet the needs. FIG. 24A shows the raw rheological data of IEIK (SEQ ID NO: 5) and IEIK (SEQ ID NO: 5) mixed with indigo carmine. FIG. 24B shows a comparison of the stiffness of IEIK (SEQ ID NO: 5) with that of IEIK (SEQ ID NO: 5) mixed with indigo carmine. FIG. 24C shows a comparison between the rigidity of RADA16 and the rigidity of RADA16 mixed with Ringer's solution.

(Example 19)
Prevention of Lung Leakage An injectable self-assembling peptide hydrogel system was used as an air sealant for pleural defects. 35cmH 2 _O or greater burst pressure (i.e., pressure air break the surface of the sealant) was achieved.

Materials and Methods Experimental Setup Pig lungs were obtained from freshly euthanized pigs. An endotracheal tube was inserted into the target lung through the trachea and main bronchus. The lungs were pressured using a manual endotracheal intubation pump. The trachea was ligated to prevent air leakage around the tube. The main bronchi of the other lung were clamped and all airflow was directed to the target lung. A manometer was used to measure the pressure of the air induced into the lungs to induce inflation.

Preparation of self-assembling peptide hydrogel self-assembling peptide hydrogel, Ac-RADARADARADARADA-NH ₂ (i.e., RADA16) (SEQ ID NO: 1), Ac-KLDLKLDLKLDL-NH 2 ( i.e., KLD12) (SEQ ID NO: 3), or Ac-IEIKIEIKIEIKI-NH ₂ is composed of (i.e., IEIK13) (SEQ ID NO: 2), RADA16 or KLD12 were mixed alone or with calcium chloride 0.250M. When alone, peptides were reconstituted in deionized water. However, if the peptide was reconstituted with a 0.250 M calcium chloride solution, the peptide was first reconstituted in deionized water and then a 0.500 M solution of calcium chloride was mixed in a 1: 1 ratio.

Creation of Pleural Defects To determine the efficacy of self-assembling peptide hydrogels, pleural defects were created by three different methods: (1) Lung and pleura were punctured using a 16 gauge needle, (2) 3.) A 5 × 5 mm area was measured, the lung and pleura were cut using one surgical scissor, and (3) a lobectomy (ie, complete resection of one lobe of the lung of interest). A 0.9% saline solution was flushed over the defect area, air was introduced, and the defect location was identified by using an endotracheal intubation pump system to observe the release of air bubbles.

Application of Self-Assembled Peptide Hydrogel Once the location of the defect was identified, the hydrogel was applied to the defect area by two different methods: (1) The hydrogel was applied locally by injecting the defect area with a syringe. And (2) the hydrogel was injected into the defect by overflow so as to cover the defect area locally with an 18 gauge needle. After a relaxation period of 2 minutes, pressure was applied through an endotracheal intubation pump until burst pressure was identified. For any additional testing, airflow to the previously tested defect was cut using a surgical clamp. FIG. 25 shows the flow of the procedure for identifying a pleural defect, inserting a needle, injecting a hydrogel, and identifying burst pressure: A) identification of a pleural defect, B) insertion of a needle into the pleural defect, C ) Injection of hydrogel (1.5% IEK13), D) Identification of bubbles indicating that burst pressure was reached.

Results Using various needle sizes, the effect of pleural defects on burst pressure was determined using 2.5% RADA16. Pleural defects were created using 14, 16, 18, and 22 gauge needles. 2.5% RADA16 was applied topically and for each defect, the burst pressure was identified using the method described above. FIG. 26 shows that using 2.5% RADA16, variations in needle gauge do not significantly alter the rupture pressure of pleural defects. The changes in burst pressure exhibited by pleural defects caused by various needle gauges were negligible.

  Using 2.5% RADA16, it was determined whether increasing the exposure time of RADA16 to a pleural defect site would increase burst pressure. A 16G pleural defect was created and 2.5% RADA16 was applied. Burst pressure was tested at various time points. FIG. 27 shows that increased exposure of RADA16 to the deficient site increased burst pressure. However, while burst pressure can generally increase with exposure time, pulmonary surgeons may only have to wait for a predetermined period of time during repair of a pleural defect, eg, about 2 minutes, before applying a sealant . The asterisk indicates the maximum possible latency assigned in a realistic surgery.

  A topically applied solution of 0.154 M NaCl (0.9% NaCl) was used to determine if the topical addition of saline solution would alter the burst pressure of 2.5% RADA16. . A 16G pleural defect was created and 2.5% RADA16 was applied topically as described above. A solution of 0.154 M NaCl was applied topically to RADA16 and the combination was gently rubbed to induce light mixing. FIG. 28 shows that adding a solution based on a salt of NaCl topically to 2.5% RADA16 and gently massaging slightly increases the burst pressure. The burst pressure was increased by topical application of saline to 2.5% RADA16.

Since the burst pressure of 2.5% RADA16 was successfully increased by topically adding a solution of NaCl to 2.5% RADA16 and gently massaging, the other salt-based solution was thoroughly mixed with RADA16. , Their mechanical properties were measured. It was theorized that gels with better mechanical properties would exhibit increased burst pressure. The results for the storage modulus (ie, mechanical strength) of 0.5% PuraMatrix (RADA 16) mixed with various concentrations of NaCl, KCl, or CaCl ₂ can be seen in FIG. Mixing RADA16 with calcium chloride showed better mechanical properties than mixing with either sodium chloride or potassium chloride.

All three hydrogels-RADA16, KLD12, and IEIK13 were used at various concentrations, and RADA16 and KLD12 were used alone and in combination with 0.250M calcium chloride. The hydrogel was applied to pleural defects as described above. FIG. 31 shows some combinations with CaCl ₂ used to determine the effectiveness of these hydrogels in preventing air leakage. FIG. 31A shows that 2.5% RADA16, 2.5% IEIK13, and 1.5% IEIK13 with 0.250 M CaCl ₂ are all 35 cm H ₂ O using the injection method to seal 16G pleural defects. Rupture pressure was exceeded. FIGS. 31B and 31C show that neither hydrogel solution achieved a burst pressure of 35 cm H ₂ O when used as a sealant for a 5 × 5 mm pleural defect or lobectomy. With respect to 16G pleural deficiency, 2.5% RADA16 + 0.25M of CaCl ₂ and 1.5% IEIK13 the best burst pressure is shown. The asterisk indicates the optimal gel for 16G pleural defects.

Conclusion Use of an injectable, self-assembling peptide hydrogel system as an air sealant is feasible for application to certain pleural defects. As determined from the experiments described herein, 2.5% RADA16, 2.5% IEIK13, and 1.5% IEIK13 hydrogels with 0.25 M CaCl ₂ burst at 35 cm H ₂ O. Exceeding pressure can be used to prevent air leaks associated with needle punctures, staples, and sutures.

Claims (23)

A group consisting of (RADA) ₄ (SEQ ID NO: 1), (IEIK) ₃ I (SEQ ID NO: 2) and (KLDL) ₃ (SEQ ID NO: 3) for use in a method of preventing bronchial anastomosis leakage in a subject. A solution comprising a self-assembling peptide selected from the group consisting of: At least a portion of the peptide, such that the peptide can exhibit a beta-sheet structure in an aqueous solution in the presence of physiological conditions at the surgical site of the lung. Amphiphilic,
The method comprising: identifying the target area where prevention of lung leakage of the bronchial anastomosis is desired; introducing a delivery device to the target area of the bronchial anastomosis of the subject; placing in the target area, the step of administering said solution through said delivery device, viewed including the step of determining step, and the bursting pressure is removed the delivery device from the target area,
The hydrogel barrier provides a burst pressure resistance of at least 35 cmH ₂ O;
solution. a) the solution is administered through an injection to the target area in an overflow to locally cover the target area; and / or b) the solution is administered in a single dose; and / or c. 2.) The solution according to claim 1, characterized in that the solution is administered post-surgery, optionally after the surgery, which may cause postoperative bronchial anastomosis lung leakage. Before SL hydrogel barrier is formed in less than about 3 minutes, a solution of claim 1.   The solution of claim 1, wherein the method further comprises adjusting a pH of the solution.   2. The solution of claim 1, wherein the solution is substantially free of cells and / or the solution is substantially free of a drug. At least one of the effective amount and the effective concentration is based in part on the size of the target area of leakage of the lung;
Optionally, said effective amount is approximately 1 mL / cm ^{2 of} target area;
The amount effective to prevent lung leakage of the bronchial anastomosis comprises a volume ranging from about 0.1 mL to about 5 mL;
The solution according to claim 1.   2. The solution according to claim 1, comprising a buffer.   The solution of claim 7, wherein the buffer comprises at least two salts.   The buffer is at a pH of 7.2 or 7.4, and / or the buffer is an alkaline buffer, and / or the solution is about 0.15 M sodium chloride, potassium chloride, magnesium chloride, 9. A solution according to claim 7 or claim 8, wherein the solution is buffered with at least one of calcium chloride. Wherein the buffer is
A salt between about 0.6M to about 1.2M, wherein the self-assembling peptide is (RADA) ₄ (SEQ ID NO: 1) or a salt between about 0.02M to about 0.04M. Wherein the self-assembling peptide is (IEIK) ₃ I (SEQ ID NO: 2) or comprises between about 0.1 M to about 0.4 M of the salt, and wherein the self-assembling peptide is (KLDL) ₃ ( SEQ ID NO: 3)
The solution according to any one of claims 7 to 9. A concentration effective to prevent leakage of the lung, including self-assembling peptide concentration in the range of from about 0.1 weight / volume (w / v)% to about 3w / v percent, of claim 1 solution.   11. The solution of claim 10, wherein the solution comprises a salt or the solution is an aqueous peptide solution. The salt is at least one cation selected from the group consisting of ammonium ion, iron ion, magnesium ion, potassium ion, pyridinium ion, quaternary ammonium ion, sodium ion, potassium ion, and calcium ion; and / or At least one selected from the group consisting of chloride ions, sulfate ions, acetate ions, carbonate ions, citrate ions, cyanide ions, fluoride ions, sulfate ions, nitrate ions, nitrite ions, and phosphate ions. containing anions, the solution according to claim 1 2.   10. The method of any of claims 7-9, wherein the solution has a salt concentration between about 0.005M to about 1M and / or the solution comprises sodium chloride, potassium chloride, calcium chloride, and sodium bicarbonate. The solution according to claim 1.   Wherein the solution further comprises a contrast agent, optionally the contrast agent comprises sulfate and sodium ions, and / or the solution further comprises at least one biologically active agent. Item 6. The solution according to Item 1. The solution has a pH of about 2.5 to about 4.0, and optionally, the solution has a pH of about 3.5, and the self-assembling peptide is (RADA) ₄ (SEQ ID NO: 1). And (KLDL) ₃ (SEQ ID NO: 3) or the solution has a pH of about 3.7 and the self-assembling peptide is (IEIK) ₃ I (SEQ ID NO: 2) in a solution according to claim 1 2 or 1 3. The solution comprises (RADA) ₄ (SEQ ID NO: 1) at a concentration of about 2.5 weight / volume (w / v) percent;
Optionally, said solution comprises a calcium chloride concentration of about 0.125 M,
Further optionally, the solution has a storage modulus of about 600 Pa.
The solution according to any one of claims 7 to 9.   2. The solution of claim 1, wherein the target area is identified by flowing the solution over the bronchial anastomotic leak and observing the release of gas bubbles.   The solution of claim 1, wherein said solution comprises calcium chloride.   Said at least 35 cmH ₂ 20. The solution of claim 19, wherein the burst pressure resistance of O is achieved for defects having a size brought up to 16G needle.   Said at least 35 cmH ₂ The solution of claim 1, wherein the burst pressure resistance of O is achieved for a defect having a size brought up to 16G needle.   Said at least 35 cmH ₂ 20. The solution of claim 19, wherein the burst pressure resistance of O is achieved for defects having a size of less than about 5 x 5 mm.   Said at least 35 cmH ₂ The solution of claim 1, wherein the burst pressure resistance of O is achieved for defects having a size of less than about 5 x 5 mm.